Solid Dosage Forms of Bendamustine

ABSTRACT

In the present invention there is provided a pharmaceutical composition in a solid dosage form suitable or oral administration, the composition comprising bendamustine or a pharmaceutically acceptable ester, salt or solvate thereof as an active ingredient, and at least one pharmaceutically acceptable excipient, which is a pharmaceutically acceptable saccharide selected from the group consisting of one or more of a monosaccharide, a disaccharide, an oligosaccharide, a cyclic oligosaccharide, a polysaccharide and a saccharide alcohol, wherein the ratio by weight of the active ingredient to the saccharide excipient(s) is in the range of 1:1-5.

The present invention relates to solid dosage forms for oraladministration comprising bendamustine or a pharmaceutically acceptableester, salt or solvate thereof.

BACKGROUND OF THE INVENTION

Bendamustine(4-[5-[bis(2-chloroethyl)amino]-1-methylbenzimidazo-2-yl]butanoic acid,a nitrogen mustard) is an alkylating agent with bifunctional alkylatingactivity. It corresponds to the following formula I:

Bendamustine appears to be free of any cross-resistance with otheralkylating agents, which offers advantages in terms of chemotherapy forpatients who have already received treatment with an alkylating agent.

Bendamustine was initially synthesized in the German Democratic Republic(GDR). The hydrochloric acid of bendamustine was the active ingredientin a commercial product available from 1971 to 1992 under the trade nameCytostasan®. Since that time, it has been marketed in Germany under thetrade name Ribomustin® and has been widely used to treat chroniclymphocytic leukemia, non-Hodgkin's lymphoma and multiple myeloma.

The marketed product contains a lyophilized powder of bendamustinehydrochloride which is reconstituted with water for injection yielding aconcentrate. This is subsequently diluted with an aqueous solution of0.9% sodium chloride resulting in the final solution for infusion. Thisfinal solution is administered to the patient by intravenous infusionover a period of about 30 to 60 minutes.

Hydrolysis of the bis-2-chloroethylamino-group of bendamustine in waterleads to reduction in potency and to impurity formation (B. Maas et al.(1994) in Pharmazie 49: 775-777). Hence administration, usually in ahospital or at least under medical supervision, must occur immediatelyafter reconstitution of the lyophilized powder. Furthermore,reconstitution has been reported to be difficult. It may require morethan 30 minutes. Further, it is burdensome and time-consuming for thehealthcare professionals responsible for reconstituting the product inthe 2 step process.

Preiss et al. (1985) in Pharmazie 40:782-784 compared thepharmacokinetics of bendamustine hydrochloride in plasma in 7 patientsafter intravenous and oral administration respectively in a dose rangingbetween 4.2-5.5 mg/kg. The intravenous infusion prepared from thecommercially available Cytostasan® product was given over 3 minutes,whereas oral medication in an equivalent dose was taken in the form ofcapsules, containing 25 mg of bendamustine hydrochloride. The number ofcapsules to be taken by the patients varied from 10-14. After oraladministration maximal plasma levels were detectable within 1 hour. Themean oral bioavailability was calculated to be 57%, ranging from 25% to94% indicating a large inter-individual variability.

Weber (1991) (Pharmazie 46(8): 589-591) investigated the bioavailabilityof bendamustine hydrochloride in B6D2F1-mice and found that theabsorption of the drug from the gastro-intestinal tract is incompleteresulting in a bio-availability of about 40% only.

US 2006/0128777 A1 describes methods for treating cancers, characterisedby death-resistant cells and bendamustine-containing compositions ingeneral. Amongst these compositions are oral solid dosage forms, whichare capsules, tablets, pills, powders or granules, wherein the activecompound may be admixed with at least one inert excipient, such assucrose, lactose or starch. However, specific compositions were notexemplified.

In view of the stability problems with the marketed i.v. formulation,once reconstituted with water, and in order to improve the patientcompliance there has been a long-felt need for a stable oral dosage-formcomprising bendamustine which is easy to administer to the patient andprovides an increased bioavailability with less variability as comparedto the known oral dosage-form.

SUMMARY OF THE INVENTION

In order to solve the above problems the present inventors have carriedout detailed investigations. They finally succeeded in obtaining thestable pharmaceutical compositions according to the invention. Thesecompositions are suitable for oral administration and comprisebendamustine or a pharmaceutically acceptable ester, salt or solvatethereof as an active ingredient, and at least one pharmaceuticallyacceptable excipient, which compositions have an improved dissolutionprofile.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the mean plasma concentration (tablets versus capsule) vs.time curve obtained after administering bendamustine hydrochloride inthe form of prior art capsules and the tablet formulations of Examples 6to 8 (Tablets 1-3) and example 9 (formulation 3) (Tablet 4) to dogs. Itis apparent from FIG. 1 that the tablet formulations provide for highermaximum concentrations of bendamustine, as compared to the prior artcapsule.

FIG. 2 shows a flow sheet of wet granulation manufacturing trials.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to a pharmaceutical composition comprisingbendamustine or a pharmaceutically acceptable ester, salt or a solvatethereof as an active ingredient and at least one pharmaceuticallyacceptable excipient selected from monosaccharides, disaccharides,oligosaccharides, cyclic oligosaccharides, a polysaccharide andsaccharide alcohols. Preferably, the ratio by weight between the activeingredient and excipient is in the range of 1 to 1-5, preferably 1 to2-5, more preferably a ratio selected from 1:5 and 1:2.

In an embodiment the present invention relates to a pharmaceuticalcomposition in a solid dosage form for oral administration, thecomposition comprising bendamustine or a pharmaceutically acceptableester, salt or solvate thereof as an active ingredient, and at least onepharmaceutically acceptable excipient, which is a pharmaceuticallyacceptable saccharide selected from the group consisting of one or moreof a monosaccharide, a disaccharide, an oligosaccharide, a cyclicoligosaccharide, a polysaccharide and a saccharide alcohol, wherein theratio by weight of the active ingredient to the excipient is in therange of 1:1.

In a further embodiment the present invention relates to apharmaceutical composition in a solid dosage form suitable for oraladministration, the composition comprising bendamustine orpharmaceutically acceptable ester, salts or solvates thereof as anactive ingredient and at least one pharmaceutically acceptable excipientwhich is a pharmaceutically acceptable saccharide selected from thegroup consisting of one or more of a monosaccharide, a disaccharide, anoligosaccharide, a cyclic oligosaccharide, a polysaccharide and asaccharide alcohol, wherein the ratio by weight of the active ingredientto the saccharide excipient(s) is in the range of 1:2-5 and whichcomposition shows a dissolution of the bendamustine of at least 60% in20 minutes, 70% in 40 minutes and 80% in 60 minutes as measured with apaddle apparatus at 50 rpm according to the European Pharmacopoeia in500 ml of a dissolution medium at a pH of 1.5.

Further preferred embodiments within the scope of the above embodimentsare pharmaceutical compositions wherein the pharmaceutically acceptablesaccharide is selected from the group consisting of one or more of amonosaccharide, a disaccharide and an oligosaccharide, wherein the ratioby weight of the active ingredient to the saccharide excipient(s) is inthe range of 1:2-5 and which composition shows a dissolution of thebendamustine of at least 60% in 20 minutes, 70% in 40 minutes and 80% in60 minutes as measured with a paddle apparatus at 50 rpm according tothe European Pharmacopoeia in 500 ml of a dissolution medium at a pH of1.5.

The present invention is based, inter alia, on the surprising discoverythat a specific and desirable dissolution profile can be reached byincorporating a certain amount of pharmaceutically acceptablesaccharides into the pharmaceutical composition.

It has been found that if a pharmaceutically acceptable saccharideselected from the group consisting of one or more of a monosaccharide, adisaccharide, an oligosaccharide, a cyclic oligosaccharide, apolysaccharide or a saccharide alcohol and preferably selected from thegroup consisting of one or more of a monosaccharide, a disaccharide andan oligosaccharide is used as an excipient in a pharmaceuticalcomposition comprising bendamustine or pharmaceutically acceptableester, salt or solvate thereof as an active ingredient, a particularlyfavourable profile of the composition as regards stability, tablettingproperties, dissolution and impurity formation is achieved. The abovesaccharides result in a composition which shows a dissolution of thebendamustine of at least 60% in 20 minutes, 70% in 40 minutes and 80% in60 minutes as measured with a paddle apparatus at 50 rpm according tothe European Pharmacopoeia in 500 ml of a dissolution medium at a pH of1.5.

Within the above scope of the invention, any combination of one or moreof a monosaccharide, a disaccharide, an oligosaccharide, a cyclicoligosaccharide, a polysaccharide and a saccharide alcohol may be used.

It has particularly been found that particular saccharides areassociated with a particularly favourable profile of a pharmaceuticalcomposition as regards stability and dissolution. Preferred saccharidesof the composition according to the present invention are dextroseanhydrous, dextrose monohydrate, lactitol monohydrate, trehalose,sorbitol, erythritol, maltose monohydrate, mannitol, lactose anhydrous,lactose monohydrate, maltitol, xylitol, sucrose, sucrose97%+maltodextrin 3%, β-cyclodextrin, D-raffinose pentahydrate,D-melezitose monohydrate and microcrystalline cellulose. Thepharmaceutical compositions according to the present invention show goodtabletting characteristics, fast dissolution and a pharmaceuticallyacceptable stability.

The above saccharides constitute preferred embodiments of the presentinvention and any combination thereof may be used. Preferably, the ratiobetween the active ingredient and the above saccharides is in the rangeof 1:1-5, preferably 1:2-5 and more preferably a ratio selected from 1:5and 1:2.

A further preferred embodiment of the invention is a pharmaceuticalcomposition in a solid dosage form for oral administration, thecomposition comprising bendamustine or a pharmaceutically acceptableester, salt or solvate thereof as an active ingredient and at least onepharmaceutically acceptable excipient selected from dextrose anhydrous,dextrose monohydrate, lactitol monohydrate, trehalose, sorbitol,erythritol, maltose monohydrate, mannitol, lactose anhydrous, lactosemonohydrate, maltitol, xylitol, sucrose, sucrose 97%+maltodextrin 3%,β-cyclodextrin, D-raffinose pentahydrate, D-melezitose monohydrate andmicrocrystalline cellulose and which composition shows a dissolution ofthe bendamustine of at least 60% in 10 minutes, 70% in 20 minutes and80% in 30 minutes.

Particularly preferred saccharides are mannitol, maltitol, erythritol,xylitol, lactose, sucrose, glucose, sorbitol, maltose, trehalose,lactitol and dextrose (anhydrous or monohydrate) and the weight ratio ofthe active ingredient to said saccharide is preferably in the range of1:2-5. Combinations of two or more saccharides within the scope of theabove saccharides are also included within the present invention.

A person skilled in the art is well in a position to select suitablecombinations within the saccharide excipients mentioned above and obtaina composition which shows a dissolution of bendamustine of at least 60%in 20 minutes, 70% in 40 minutes and 80% in 60 minutes as measured witha paddle apparatus at 50 rpm according to the European Pharmacopoeia in500 ml of a dissolution medium at a pH of 1.5.

In a preferred embodiment the composition is in the form of a tablet, agranulate, or a pill.

A preferred dosage form is a tablet. The term tablet also comprisesfast-disintegrating tablets, amongst which are dispersible tablets andeffervescent tablets.

The most commonly used methods of tablet preparation are directcompression, dry granulation and wet granulation. Direct compressioninvolves compressing a mixture containing the active ingredient(s) andthe excipient(s) on a tablet press (L. Lachman et al., in: The Theoryand Practice of Industrial Pharmacy, 3rd ed., 1986). The mixture to becompressed must possess both good flow and compression properties inorder to produce tablets having a uniform content of the activeingredient(s). Good flow properties cannot always be achieved by addingappropriate excipients, such as lubricants, anti-adhesive agents andflow-promoters to the mixture. Hence frequently the mixture isgranulated prior to compression.

Granulation is a process by which sphere-like or regularly shapedaggregates called granules are formed out of the powder mixture. Thiscan be achieved by dry granulation methods and wet granulation methods.Granulation is also used for converting a mixture of powders with poorcohesion into aggregates, which when compressed result in tablets thathave good cohesion properties.

In the case of fast-disintegrating tablets, the active ingredient(s),optionally in admixture with one or more excipients, is (are)advantageously provided with a coating in order to mask the taste ofsuch ingredient(s) and/or to protect the same against possible harmfuleffects by light and/or moisture and in the case of bendamustine toprotect the mucosa in the mouth against the harmful effects exerted bythe active compound. For that purpose a granulate preferably is preparedand processed as further outlined below.

The expression “granulate” refers to aggregates of particles, sometimescalled granules. A granulate in general is prepared by compaction and/orcompression techniques (dry granulation) or by wet granulationtechniques, using a liquid in which optionally a wet granulation bindingagent is dissolved (Remington's Pharmaceutical Sciences 18th ed. 1990,page 1641). Wet granulation techniques also include extrusiontechniques. Accordingly the term granulate also comprises pellets,spherules, and extrudates, of which pellets preferably are used asexamples of a granulate.

A pellet may be described as a small particle of approximately 1.0-1.6mm in diameter and having a certain density, which particle is preparedby application of the pharmaceutical processes of extrusion andspheronisation to powder mixtures.

The active ingredient(s), optionally in admixture with one or moreexcipients, may be advantageously provided with a coating in order tomask the taste of such ingredient and/or to protect the same againstpossible harmful effects by light and/or moisture and/or to protect themucosa in the mouth against the harmful effects exerted by the activecompound.

Pills are small, round solid dosage forms, prepared by adding the activeingredient to a doughy mixture of triglycerides. The mixture is rolledinto a long string, which is then cut into pieces and rolled (J. T.Carstensen: Pharmaceutical principles of solid dosage forms, 1993,Technomic Publishing Company, Inc. page 63).

Preferably the dosage forms according to the invention are prepared bydry compaction techniques. Suitable techniques are for example describedin Remington's Pharmaceutical Science 18th. ed. 1990, page 1644. Theycomprise dry granulation, roller compaction and direct compression. Whentablets are prepared by these techniques, it is even more advantageousto use direct compression.

The dosage forms according to the present invention are preferablyprovided with a coating. The coating has different purposes: it mayserve for masking the taste of the active ingredient(s) used in thecomposition, whilst at the same time it is protecting the activeingredient against possible harmful effects by light and/or moisturesuch as oxidation, degradation, etc. Furthermore, the coating layer mayprevent the subject from damage of the oral mucosa by the activeingredient.

The coating layer can be applied to the dosage forms by techniqueswell-known in the art such as spray-coating and microencapsulation. Fortablets it can be in the form of a film-coating, a saccharide-coating ora compression coating. Preferably a film-coating process is used(Remington's Pharmaceutical Sciences 18th ed. 1990, page 1666). In casean active ingredient requires the application of a coating forfast-disintegrating tablets the individual granules can suitably beprovided with a coating prior to compression into tablets.

The expression “pharmaceutically acceptable ester thereof” describes anypharmaceutically acceptable ester of bendamustine, such as esters withalkyl alcohols and saccharide alcohols. Examples of the alkyl alcoholsare C1-6-alkyl alcohols such as methanol, ethanol, propanol,isopropanol, butanol and tert-butanol. Examples of the saccharidealcohols are mannitol, maltitol, sorbitol, erythritol, glycol, glycerol,arabitol, xylitol and lactitol. Preferred examples of the bendamustineesters are the ethyl ester, the isopropyl ester, the mannitol ester andthe sorbitol ester, most preferred is the ethylester thereof.

The expression “pharmaceutically acceptable salt thereof” describes anypharmaceutically acceptable salt of bendamustine that administered to apatient (directly or indirectly) provides bendamustine. This termfurther comprises the pharmaceutically acceptable salt of a bendamustineester. Nevertheless, it will be considered that the pharmaceuticallynon-acceptable salts also are included within the limits of thisinvention since these compounds can be useful in the preparation ofpharmaceutically acceptable salts. For example, pharmaceuticallyacceptable salts of bendamustine are synthesized from the correspondingcompound that contains an acid or basic group, by conventional chemicalmethods. Generally, these salts are, for example, prepared by means ofthe reaction of free acidic or basic forms of these compounds in astoichiometric amount with a corresponding base or acid in water or anorganic solvent or a mixture of both. Nonaqueous media like ether, ethylacetate, isopropanol or acetonitrile are generally preferred. Examplesof acids which may be used for the salt formation of pharmaceuticallyacceptable salts of bendamustine include inorganic acids such ashydrochloride, hydrobromide, hydriodide, sulphuric, nitric, andphosphoric acids, and organic acids such as acetic, maleic, fumaric,citric, oxalic, succinic, tartaric, malic, lactic, methylsulphonic andp-toluenesulphonic acids. Pharmaceutically acceptable salts ofbendamustine may be derived from either inorganic or organic bases toyield ammonium salts; alkali metal salts (lithium, sodium, potassium,etc.), alkaline earth salts like calcium or magnesium, aluminium salts,lower alkylamine salts like methylamine or ethylamine salts, loweralkyldiamine salts like ethylenediamine salts, ethanolamine,N,N-dialkyleneethanolamine, triethanolamine, and glucamine salts, aswell as basic salts of amino acids. Especially preferred are acid saltsprepared from the hydrochloride, the hydrobromide, and the hydroiodide,whereas the hydrochloride salt is the most preferred pharmaceuticallyacceptable salt of bendamustine. The pharmaceutically acceptable saltsare produced by conventional techniques well-known in the art.

The expression “pharmaceutically acceptable solvate thereof” describesany pharmaceutically acceptable solvate that, administered to a patient(directly or indirectly) provides bendamustine. This term furthercomprises the pharmaceutically acceptable solvate of a bendamustineester. Preferably, the solvate is a hydrate, a solvate with an alcoholsuch as methanol, ethanol, propanol, or isopropanol, a solvate with anester such as ethyl acetate, a solvate with an ether such as methylether, ethyl ether or THF (tetrahydrofuran) or a solvate with DMF(dimethylformamide), of which a hydrate or a solvate with an alcoholsuch as ethanol is more preferred. A solvent for constituting thesolvate is preferably a pharmaceutically acceptable solvent.

It is especially preferred that the active ingredient in the invention'scompositions is bendamustine or a pharmaceutically acceptable saltthereof. It is most preferred that the active ingredient is bendamustinehydrochloride.

The dose of the active ingredient in the pharmaceutical composition mayreadily be determined by the skilled artisan depending on the patient'scondition, sex, body weight, body surface area, or age, especiallydepending on the patient's body weight and body surface area and rangesfrom 10 to 1000 mg. It is preferred that the daily dosage ranges fromabout 50 to about 1000 mg, preferably from about 100 to about 500 mg ofthe active ingredient. The daily dosage may be taken as a single dose oras a multiple dose such as twice or three-times daily, most preferablyas a single daily dose. The daily dose may be taken once a week orseveral times a week. The dosage form may contain the amount of a singledaily dose or parts thereof. It is preferred that the dosage form of thepresent invention comprises about 10 to about 1000 mg, preferably about25 to about 600 mg, more preferably about 50 to about 200 mg and mostpreferably about 100 mg of the active ingredient.

The saccharides are present in the compositions according to theinvention in a substantial amount, preferably in an amount ranging from2 to 5 times the weight of the active substance. The saccharides whenincorporated into the compositions of the present invention, have shownto have a positive effect on the stability of the active compound. Inaddition to that it was surprisingly found that these excipients resultin an increased bio-availability of the active compound, in particularbendamustine hydrochloride, when compared to the reference capsule.

Preferred examples of the saccharides include mannitol, maltitol,erythritol, xylitol, lactose, sucrose, glucose, sorbitol, maltose,trehalose, lactitol and dextrose (anhydrous or monohydrate).

In addition to these saccharide excipients the pharmaceuticalcomposition according to the present invention may comprise furtherexcipients as described in more detail below for lubricants, glidants,fillers (or diluents), binders and disintegrants.

Lubricants are substances which may have one or more of the followingfunctions in pharmaceutical compositions and especially in tabletmanufacture: preventing adhesion of the tablet material to the surfaceof parts of the tabletting machine (hopper, dies and punches), reducinginterparticle friction, facilitating ejection of the tablets from thedies and improving the flow rate of the mixture (to be tabletted). Saidlubricant is typically selected from a group consisting of stearic acid,salts or esters of stearic acid, hydrogenated vegetable oils, magnesiumoxide, polyethylene glycol, sodium lauryl sulphate and talc, andmixtures thereof. Preferably said lubricant is selected from magnesiumstearate, calcium stearate, zinc stearate, glyceryl palmitostearate andsodium stearyl fumarate, and mixtures thereof. Stearic acid is the mostpreferred alternative.

The term glidant in this application is to be understood as a substancewhich improves the flow characteristics of the mixture to be tabletted.With respect to glidants, any suitable glidant such as talc, silicondioxide and silicagel (Cab-O—Sil®, Syloid®, Aerosil®), starch andcalcium silicate may be used. Typically, silicon dioxide is used.

Generally, the terms filler (or diluent) represent excipients which areused to increase the bulk of the materials to be tabletted. Thisincrease in size improves the handling of the solid compositions.Fillers are usually necessary if the dose of drug per solid compositionis low and the solid composition would otherwise be too small. Examplesof suitable fillers are lactose, sucrose, mannitol, sorbitol,saccharose, starch, pregelatinized starch, microcrystalline cellulose,powdered cellulose, calcium hydrogen phosphate, calcium carbonate andany combinations thereof. In a preferred embodiment the filler isselected from the group consisting of lactose, starch, microcrystallinecellulose, microfine cellulose and any combinations thereof, mostpreferably anhydrous lactose and microcrystalline cellulose.

Generally, the term binder is used for agents that impart cohesivenessto the pharmaceutical formulation, which cohesiveness ensures that thecomposition remains intact especially in case of tablets aftercompression. Dependent on the compaction technique used (directcompression, dry granulation or wet granulation) different binders areused. For dry compaction techniques (direct compression and drygranulation) suitable binders are lactose, sucrose, mannitol, sorbitol,saccharose, starch, pregelatinized starch, microcrystalline cellulose,powdered cellulose, calcium hydrogen phosphate, calcium carbonate andany combinations thereof. In a preferred embodiment the binder isselected from the group consisting of lactose, starch, microcrystallinecellulose, microfine cellulose and any combinations thereof, mostpreferably anhydrous lactose and microcrystalline cellulose. In wetgranulation processes binders can be used both as a solution and in adry form. As suitable binders, there may be mentioned, for example,polyvinylpyrrolidone, dispersible cellulose, hydroxypropyl cellulose,hydroxypropylmethyl cellulose, methylcellulose, starch, pregelatinizedstarch, partly pregelatinized starch, gum arabic, dextrin, pullulan andthe like. Among these binders, dispersible cellulose,polyvinylpyrrolidone, hydroxypropyl cellulose and hydroxypropylmethylcellulose are more preferred.

A disintegrant can be included in a pharmaceutical composition andespecially a tablet composition to facilitate its breakup ordisintegration after the tablet comes into contact with a physiologicalaqueous liquid. When the tablet is swallowed, the disintegrant often isresponsible for the quick disintegration of the tablet when it comesinto contact with body fluids, such as saliva, gastric and intestinalfluids. Materials serving as disintegrants have been classifiedchemically as starches, celluloses, cross-linked polymers, etc. As aresult of investigations concerning the disintegrator species to be usedin the practice of this invention and the level of addition thereof, itwas found that starch, a modified starch such as sodium starch glycolate(Primojel®), sodium carboxymethyl cellulose, crosslinkedcarboxymethylcellulose sodium (Ac-Di-Sol®), cross-linkedpolyvinylpyrrolidone, polacrilin potassium (Amberlite® IRP88) andlow-substituted hydroxypropyl cellulose can produce a very gooddisintegrating effect.

The stability of an aqueous solution of bendamustine is stronglyinfluenced by the pH. A significant hydrolytic decomposition of thiscompound is observed at pH values higher than about 5. At pH>5, thedecomposition proceeds rapidly and the resulting content of by-productsis high in this pH range. The main hydrolysis products are4-[5-[(2-Chloroethyl)-(2-hydroxy-ethyl)amino]-1-methyl-benzimidazo-2-yl]-butanoicacid (HP1),4-[5-[Bis(2-hydroxyethyl)amino]-1-methyl-benzimidazo-2-yl]-butanoic acid(HP2) and 4-(5-Morpholino-1-methylbenzimidazol-2-yl)-butanoic acid(HP3):

Absorption of an orally administered drug usually happens in thestomach, the small intestine and/or the large intestine. The pH in thestomach is about 1 to 3.5, in the small intestine about 6.5 to 7.6, andin the large intestine about 7.5 to 8.0. Accordingly, for a compoundlike bendamustine which is prone to degradation in aqueous environmentswith a pH higher than 5, it is highly preferable that it is absorbed inthe stomach, and does not pass through to the small or even the largeintestine, in order to avoid decomposition. Hence there is a need for apharmaceutical composition from which the bendamustine is absorbedcompletely or at least to a high extent in the stomach, thereby avoidingor reducing the degradation of the bendamustine in the small or largeintestine.

Applicant has found that surprisingly it is possible to solve thisproblem by using the present pharmaceutical compositions and, inparticular, the pharmaceutical composition with the above preferredsaccharides. These compositions containing bendamustine show a fastdissolution, and in particular a dissolution of the bendamustine of atleast 60% in 20, preferably 10 minutes, 70% in 40, preferably 20 minutesand 80% in 60, preferably 30 minutes, and most preferably of at least75% in 10 minutes, 85% in 20 minutes and 90% in 30 minutes, as measuredwith a paddle apparatus at 50 rpm according to the EuropeanPharmacopoeia in an artificial gastric fluid. The artificial gastricfluid as used herein refers to a solution prepared by dissolving 2 g ofsodium chloride in 1000 ml of water and then adjusting the pH to1.5±0.05 with 5 N hydrochloric acid.

The total time of a drug to pass the stomach to the small intestine isbetween about 20 minutes to 5 hours, usually between about 30 minutes to3 hours. Thus pharmaceutical compositions according to this inventionadvantageously should reduce the degradation of bendamustine in thepatient since the bendamustine is released and dissolved to a majorextent while in the stomach, thus resulting in an improvedbioavailability of the bendamustine containing compositions according tothe invention.

In a further aspect of this invention the pharmaceutical compositions ina solid dosage form may be used for the treatment, induction, salvagetherapy, conditioning prior to stem cell transplantation, maintenancetherapy, treatment of residual disease of a medical condition in a humanor animal, preferably a human, which medical condition is selected fromchronic lymphocytic leukemia (CLL), acute lymphocytic leukaemia (ALL),chronic myelocytic leukaemia (CML), acute myelocytic leukaemia (AML),Hodgkin's disease, non-Hodgkin's lymphoma (NHL), multiple myeloma,breast cancer, ovarian cancer, small cell lung cancer, non-small celllung cancer, and an autoimmune disease.

The present invention also comprises a method of treatment of a medicalcondition selected from chronic lymphocytic leukemia, acute lymphocyticleukaemia, chronic myelocytic leukaemia acute myelocytic leukaemia,Hodgkin's disease, non-Hodgkin's lymphoma, multiple myeloma, breastcancer, ovaryian cancer, small cell lung cancer, non-small cell lungcancer, and an autoimmune disease, in a human or animal body comprisingadministering to the human or animal body in need thereof an effectiveamount of the pharmaceutical preparation of this invention. Preferablythe medical condition is non-Hodgkin's lymphoma.

In another aspect the of this invention the pharmaceutical compositionmay be administered in combination with at least one further activeagent, wherein said further active agent is given prior, concurrently,or subsequently to the administration of the pharmaceutical composition.This at least one further active agent is preferably an antibodyspecific for CD20 (such as rituximab or ofatumumab), an anthracyclinderivative (such as doxorubicin or daunorubicin), a vinca alkaloid (suchas vincristine), a platin derivative (such as cisplatin or carboplatin),daporinad (FK866), YM155, thalidomide and analogues thereof (such asthalidomide or lenalidomide), or a proteasome inhibitor (such asbortezumib).

The pharmaceutical composition of this invention may also beadministered in combination with at least one corticosteroid, whereinsaid corticosteroid is given prior, concurrently, or subsequently to theadministration of the pharmaceutical composition. Examples of thecorticosteroids are prednisone, prednisolone or dexamthasone.

The following examples further illustrate the invention. It will beapparent to the skilled person that these examples are solely forillustrative purposes and must not be considered to limit the invention.

EXAMPLES 1. Compatibility Tests Example 1a

For compatibility testing mixtures containing bendamustine hydrochlorideand an excipient at a ratio of 1:1 (m/m) were prepared. The excipientswere selected from mannitol and lactose. After preparation the mixtureswere packed in clear glass HPLC-Vials (6 ml) Agilent and stored atdifferent storage conditions as shown in Table 1 below. At defined timepoints samples were removed from storage and tested for purity (HPLC;column: Zorbax Bonus-RP, 5 μm; temperature of column oven: 30° C.;temperature of autosampler: 5° C.; detector: 254 nm) and appearance.

TABLE 1 Storage Conditions Bendamustine hydrochloride and excipients fororal formulation Tested time points T = 1 Storage condition T = 0 month(1) 50° C., Vials closed n = 2 n = 1 (2) 70° C., Vials closed* n = 2 n =2 (3) 40° C./75% r.h., Vials open** n = 2 n = 2 *stored at 50° C. forone month before storage at 70° C. **stored at 25° C./60% r.h. for onemonth before storage at 40° C./75%

In all these mixtures, the bendamustine hydrochloride content (measuredby HPLC) barely changed and always remained above 99% for all threestorage conditions. The hydrolysis product HP 1 was barely detectable(Area %<0.2) for all three storage conditions.

Appearance tests of the named bendamustine hydrochloride mixtures werecarried out with the naked eye. All the investigated mixtures compliedwith the specifications and gave white to off-white powder bothimmediately after preparation and after one month of storage under allthree storage conditions.

Example 1b

For further compatibility testing in accordance with the methods ofexample 1a, mixtures containing bendamustine hydrochloride and anexcipient at a ratio of 1:1 (m/m) were prepared. The excipients wereselected from Opadry®, Eudragit® E PO, sodium carboxymethylcellulose(Avicel® RC 591) and cross-linked polyvinylpyrrolidone (Crospovidone).

In the case of Eudragit® E PO the initial amounts of the impurities HP1(hydrolysis product) and BM1DIMER were significantly increased (HP 1:1.5%, BM1DIMER: 1%) but during storage a decrease of these impuritiescould be detected at all storage conditions independent of the influenceof humidity. In the case of cross-linked polyvinylpyrrolidone asignificant increase of HP 1 from 0.1% to 0.4% could be detected at thestorage condition 40°/75% R.H./vials open. At all other storageconditions (vials closed) no increase of HP1 could be detected.

The appearance of the mixtures containing Eudragit® E PO andcross-linked polyvinylpyrrolidone changed at the storage condition 70°C./vials closed. Both mixtures got lightly sticky. Additionally thecolour of the mixture with cross-linked polyvinylpyrrolidone changedfrom white too cream-coloured.

In case of the mixtures containing Opadry® and Avicel® RC591 the colouralso changed to cream-coloured at the storage condition 70° C./vialsclosed.

2. Tablet Formulations Example 2

253 g of a mixture comprising mannitol as the main excipient andmicrocrystalline cellulose, Ac-Di-Sol®, colloidal silicon dioxide, talcand stearic acid in the relative quantities mentioned in the followingtable 2a was prepared by mixing in a 1 liter cube blender (Erweka) for15 minutes. Thereafter 10.612 g of the mixture and 3.0 g of bendamustinehydrochloride were sieved through a 0.425 mm sieve and then transferredinto a Turbula mixer T2A, equipped with a glass vial of 50 ml andsubsequently mixed for 10 minutes at 60 rpm.

From this mixture round tablets were compressed having the followingcharacteristics:

Mean value diameter: 9.1 mm; mean value mass: 247.7 mg; mean valuehardness: 81N.

TABLE 2a Tablet Component mg/dosage-form Relative Content % bendamustinehydrochloride 55.1 22.04 Mannitol 141.4 56.56 Microcrystalline cellulose25.0 10.00 (Avicel ® PH112) Ac-Di-Sol ® 12.5 5.00 Colloidal silicondioxide 1.0 0.40 (Aerosil ® 200) Talc 12.5 5.00 Stearic acid 2.5 1.00

Tablets were stored at 40° C./75% RH (glass vial open) or 50° C. (glassvial closed). The amount of bendamustine hydrochloride as well as ofrelated substances, like degradation products, by-products of synthesiswere measured with HPLC (column: Zorbax Bonus-RP, 5 μm; temperature ofcolumn oven: 30° C.; temperature of autosampler: 5° C.; detector: 254nm). The results are shown in Table 2b.

TABLE 2b Related substances and assay of bendamustine HCl (residualcontent) Bendamustine HCl [% area] Storage Related T = 1 T = 1 conditionsubstances*¹ T = 0*² month T = 0 month 40° C./75% RH HP1 0.13 0.22 99.6099.13 (open vial) NP1 0.02 0.02 BM1Dimer 0.06 0.25 BM1EE 0.13 0.12 HP2n.d. 0.13 HP3 n.d. 0.03 50° C. HP1 0.13 0.53 99.60 98.94 (closed vial)NP1 0.02 0.02 BM1Dimer 0.06 0.14 BM1EE 0.13 0.11 HP2 n.d. 0.05 HP3 n.d.n.d. *¹NP1:4-[6-(2-Chloroethyl)-3,6,7,8-tetra-hydro-3-methyl-imidazo[4,5-h]-[1,4]benzothiazin-2-yl]butanoic acid BM1Dimer:4-{5-[N-(2-Chloroethyl)-N-(2-{4-[5-bis(2-chloroethyl)amino-1-methylbenzimidazol-2-yl]butanoyloxy}ethyl)amino]-1-methylbenzimidazol-2-yl} butanoic acid BM1EE:4-[5-[Bis(2-chloroethyl)amino]-1-methyl-benzimidazo-2-yl] butanoic ethylester *²n.d.: not detectable, i.e. beyond detection limit (areapercentage less than 0.05%)

Example 3

A mixture and tablets were prepared in the same way as described inExample 2, but using the compounds and relative amounts as indicated inthe following Table 3a.

The tablets had the following characteristics:

Mean value diameter: 9.1 mm; mean value mass: 248.9 mg.

TABLE 3a Tablet Component mg/dosage-form Relative Content % bendamustinehydrochloride 55.1 22.04 Lactose anhydrous 141.4 56.56 Microcrystallinecellulose 25.0 10.00 (Avicel ® PH112) Ac-Di-Sol ® 12.5 5.00 Colloidalsilicon dioxide 1.0 0.40 (Aerosil ® 200) Talc 12.5 5.00 Stearic acid 2.51.00

Tablets were stored at 40° C./75% RH (glass vial open) or 50° C. (glassvial closed). The amount of bendamustine hydrochloride and of relatedsubstances was measured with HPLC as mentioned above. The results areshown in Table 3b:

TABLE 3b Related substances and assay of bendamustine HCl (residualcontent) Bendamustine HCl [% area] Storage Related T = 1 T = 1 conditionsubstances T = 0 month T = 0 month 40° C./75% RH HP1 0.12 0.22 99.6099.13 (open vial) NP1 0.02 0.02 BM1Dimer 0.06 0.26 BM1EE 0.13 0.13 HP2n.d. 0.11 HP3 n.d. 0.03 50° C. HP1 0.12 0.57 99.61 98.88 (closed vial)NP1 0.02 0.02 BM1Dimer 0.06 0.13 BM1EE 0.13 0.11 HP2 n.d. 0.05 HP3 n.d.n.d.

Example 4

Tablets were prepared in the same way as described in Example 2, butusing the compounds and relative amounts as indicated in the followingTable 4a.

The tablets had the following characteristics:

Mean value diameter: 9.1 mm; mean value mass: 247.8 mg.

TABLE 4a Tablet Component mg/dosage-form Relative Content % Bendamustinehydrochloride 55.1 22.04 Lactose anhydrous 145.15 58.06 Microcrystallinecellulose 31.25 12.50 (Avicel ® PH112) Ac-Di-Sol ® 12.5 5.00 Colloidalsilicon dioxide 1.0 0.40 (Aerosil ® 200) Magnesium stearate 2.5 1.00Ascorbic acid 2.5 1.00

Tablets were stored at 40° C./75% RH (glass vial open) or 50° C. (glassvial closed). The amount of bendamustine hydrochloride and of relatedsubstances was measured with HPLC as mentioned above. The results areshown in Table 4b:

TABLE 4b Related substances and assay of bendamustine HCl (residualcontent) Bendamustine HCl [% area] Storage Related T = 1 T = 1 conditionsubstances T = 0 month T = 0 month 40° C./75% RH HP1 0.13 0.24 99.5899.05 (open vial) NP1 0.02 0.02 BM1Dimer 0.06 0.27 BM1EE 0.14 0.13 HP2n.d. 0.13 HP3 n.d. 0.06 50° C. HP1 0.13 0.63 99.58 98.32 (closed vial)NP1 0.02 0.02 BM1Dimer 0.06 0.18 BM1EE 0.14 0.12 HP2 n.d. n.d. HP3 n.d.n.d.

Prior Art Reference Example

20.0±1 mg of bendamustine hydrochloride were weighed into the body of anempty hard gelatine capsule, and put into a clear glass HPLC vial (6 ml)of Agilent. Capsules were closed by placing the cap on top of the bodyand slight pushing. Capsules were stored at 40° C./75% RH (glass vialopen) or 50° C. (glass vial closed). The amount of bendamustinehydrochloride and of related substances was measured with HPLC asmentioned above.

The results are shown in Table 5:

TABLE 5 Related substances and assay of bendamustine HCl (residualcontent) Bendamustine HCl [% area] Storage Related T = 1 T = 1 conditionsubstances T = 0 month T = 0 month 40° C./75% RH HP1 0.10 0.45 99.6498.83 (open vial) NP1 0.02 0.02 BM1Dimer 0.06 0.42 BM1EE 0.13 0.11 HP2n.d. n.d. HP3 n.d. n.d. 50° C. HP1 0.10 1.46 99.64 97.51 (closed vial)NP1 0.02 0.02 BM1Dimer 0.06 0.24 BM1EE 0.13 0.12 HP2 n.d. n.d. HP3 n.d.n.d.

As is immediately apparent, the capsule formulations were a lot lessstable than the tablet formulations according to the invention althoughthe capsule formulations were prepared from pure bendamustinehydrochloride without any further processing steps. Both at 40° C./75%RH (glass vial open) and 50° C. (closed vial) more degradation productsare formed within one month of storage. In the case of open vial with40° C. and 75% RH (relative humidity) the amount of hydrolysis productHP1 is increased by a factor of 4 after one month of storage. For theclosed vials the HP1 content is even higher, which might be due toreaction with the capsules. Summarising, tablets provide a much morestable solid dosage form than the capsules.

Example 5

8.0 g of hydroxypropylmethyl cellulose and 1.5 g PEG 6000 are dissolvedin 88.5 g purified water. Thereafter 2.0 g yellow ferric oxide and 0.5 gtitanium oxide are dispersed therein yielding a coating liquid. Tabletsas obtained in Example 2 are coated with 3% of this solution per tabletmass using a film coating device.

Example 6

TABLE 6a Coated Tablet Component mg/dosage-form Relative Content %Tablet cores bendamustine hydrochloride 55.1 21.09 Mannitol 141.4 54.11Microcrystalline cellulose 25.0 9.57 (Avicel ® PH101) Crosscarmellosesodium 12.5 4.78 (Ac-Di-Sol ®) Colloidal silicon dioxide 1.0 0.38(Aerosil ® 200) Talc 18.8 7.19 Stearic acid 7.5 2.87 Sum 261.3 100Film-coating Opadry ® 12.5 10 Purified water — 90 Target mass gain 12.5100 (mg/tablet)/Sum Manufacturing method for 1000 tablets

All tablet-core components except for colloidal silicon dioxide andstearic acid were loaded into a Somakon vessel (5 L). Bendamustine wasadded and blending was conducted for 4 minutes at 1000 rpm (wiper 10rpm). The resulting blend was sieved through a 0.5 mm sieve. The vesselwas reloaded with the blend and colloidal silicon dioxide was added.Blending was conducted for 2 minutes at the afore-mentioned conditions.Thereafter stearic acid was added and blending was continued for 1minute. The blend was subsequently sieved through a 0.5 mm sieve,reloaded into the vessel and blended for another 30 seconds, all at thesame conditions.

From this mixture round tablets were compressed having the followingcharacteristics:

Mean value diameter: 9.5 mm; mean value mass: 254.6 mg (begin)—257.2 mg(end); friability 0.1%; mean value hardness: 122N (begin)—128 (end).

The tablets were subsequently film-coated with the Opadry® dispersionuntil a mass increase of 5% had been achieved.

The mean mass of the film-coated tablets was 268.4 mg.

Both the tablet cores and film-coated tablets were stored at 40° C./75%RH in closed amber glass vials. The amount of bendamustine hydrochlorideas well as of related substances, like degradation products, by-productsof synthesis were measured with HPLC as mentioned above. The results areshown in Tables 6b.1 and 6b.2.

TABLE 6b.1 Related substances and assay of bendamustine HCl (residualcontent) in tablet cores Bendamustine HCl [% area] Storage Related T = 2T = 2 condition substances T = 0 months T = 0 months 40° C./75% RH HP10.15 0.13 99.49 99.49 (closed vials) NP1 n.d. n.d. BM1Dimer 0.09 0.17BM1EE 0.15 0.13 Unid RRT 0.08 0.05 0.69*³ *³Unidentified compound peakat relative retention time of 0.69 as compared to main peak

TABLE 6b.2 Related substances and assay of bendamustine HCl (residualcontent) in coated tablet Bendamustine HCl [% area] Storage Related T =2 T = 2 condition substances T = 0 months T = 0 months 40° C./75% RH HP10.16 0.17 99.46 99.29 (closed vials) HP2 n.d. 0.08 HP3 n.d. <0.05 NP1n.d. n.d. BM1Dimer 0.09 0.18 BM1EE 0.15 0.14 Unid RRT 0.10 0.05 0.69

Example 7

TABLE 7a Tablet Component mg/dosage-form Relative Content % Tablet coresbendamustine hydrochloride 55.1 21.09 Lactose anhydrous 141.4 54.11Microcrystalline cellulose 25.0 9.57 (Avicel ® PH112) Crosscarmellosesodium 12.5 4.78 (Ac-Di-Sol ®) Colloidal silicon dioxide 1.0 0.38(Aerosil ® 200) Talc 18.8 7.19 Stearic acid 7.5 2.87 Sum 261.3 100Film-coating Eudragit ® E PO 7.5 7.5 Sodium laurylsulphate 0.8 0.8Stearic acid 1.2 1.2 Iron oxide 1.0 1.0 Titanium dioxide 1.0 1.0 Talc3.5 3.5 Purified water — 85.0 Target mass gain 15.0 100.0(mg/tablet)/Sum Manufacturing method for 1000 tablets

All tablet-core components except for colloidal silicon dioxide andstearic acid were loaded into a Somakon vessel (5 L). Bendamustine wasadded and blending was conducted for 4 minutes at 1000 rpm (wiper 10rpm). The resulting blend was sieved through a 0.5 mm sieve. The vesselwas reloaded with the blend and colloidal silicon dioxide was added.Blending was conducted for 2 minutes at the afore-mentioned conditions.Thereafter stearic acid was added and blending was continued for 1minute. The blend was subsequently sieved through a 0.5 mm sieve,reloaded into the vessel and blended for another 30 seconds, all at thesame conditions.

From this mixture round tablets were compressed having the followingcharacteristics:

mean value diameter: 9.5 mm; mean value mass: 262.4 mg (begin)—254.4 mg(end); friability: 0.1% (begin)—0.2% (end); mean hardness value: 98N(begin)—91N (end). The tablets were subsequently film-coated with theEudragit® dispersion until a mass increase of 3% had been achieved.

The mean mass of the film-coated tablets was 273.5 mg.

Both the tablet cores and the film-coated tablets were stored at 40°C./75% RH in closed amber glass vials. The amount of bendamustinehydrochloride and of related substances was measured with HPLC, asmentioned above. The results are shown in Tables 7b.1 and 7.b2:

TABLE 7b.1 Related substances and assay of bendamustine HCl (residualcontent) in tablet core Bendamustine HCl [% area] Storage Related T = 2T = 2 condition substances T = 0 months T = 0 months 40° C./75% RH HP10.17 0.12 99.50 99.55 (closed vials) NP1 n.d. n.d. BM1Dimer 0.09 0.14BM1EE 0.15 0.14 Unid RRT 0.06 0.05 0.69

TABLE 7b.2 Related substances and assay of bendamustine HCl (residualcontent) in coated tablet Bendamustine HCl [% area] Storage Related T =2 T = 2 condition substances T = 0 months T = 0 months 40° C./75% RH HP10.18 0.20 99.43 98.93 (closed vials) HP2 n.d. 0.35 HP3 n.d. 0.07 NP1n.d. n.d. BM1Dimer 0.12 0.20 BM1EE 0.15 0.13 Unid RRT 0.05 <0.05 0.69

Example 8

TABLE 8a Tablet Component mg/dosage-form Relative Content % Tablet coresBendamustine hydrochloride 55.1 22.04 Lactose anhydrous 145.15 58.06Microcrystalline cellulose 31.25 12.50 (Avicel ® PH112) Ac-Di-Sol ® 12.55.00 Colloidal silicon dioxide 1.0 0.40 (Aerosil ® 200) Magnesiumstearate 2.5 1.00 Ascorbic acid 2.5 1.00 Sum 250 100.0 Film-coatingEudragit ® E PO 7.5 7.5 Sodium laurylsulphate 0.8 0.8 Stearic acid 1.21.2 Iron oxide 1.0 1.0 Titanium dioxide 1.0 1.0 Talc 3.5 3.5 Purifiedwater — 85.0 Target mass gain 15.0 100.0 (mg/tablet)/Sum Manufacturingmethod for 1000 tablets

All tablet-core components except for colloidal silicon dioxide andstearic acid were loaded into a Somakon vessel (2.5 L). Bendamustine wasadded and blending was conducted for 4 minutes at 1000 rpm (wiper 10rpm). The resulting blend was sieved through a 0.5 mm sieve. The vesselwas reloaded with the blend and colloidal silicon dioxide was added.Blending was conducted for 2 minutes at the afore-mentioned conditions.Thereafter stearic acid was added and blending was continued for 1minute. The blend was subsequently sieved through a 0.5 mm sieve,reloaded into the vessel and blended for another 30 seconds, all at thesame conditions.

From this mixture round tablets were compressed having the followingcharacteristics:

Mean value diameter: 9.5 mm; mean value mass: 252.2 mg (begin)—250.7 mg(end); friability: 0.1% (begin)—0.2% (end); mean hardness value: 65N(begin)—73N (end).

The tablets were subsequently film-coated with the Eudragit® dispersionuntil a mass increase of 3% had been achieved.

The mean mass of the film-coated tablets was 253.6 mg.

Both the tablet cores and the film-coated tablets were stored at 40°C./75% RH in closed amber glass vials. The amount of bendamustinehydrochloride and of related substances was measured with HPLC, asdescribed above. The results are shown in Tables 8b.1 and 8b.2:

TABLE 8b.1 Related substances and assay of bendamustine HCl (residualcontent) in tablet core Bendamustine HCl [% area] Storage Related T = 2T = 2 condition substances T = 0 months T = 0 months 40° C./75% RH HP10.17 0.14 99.47 99.45 (closed vials) HP3 n.d. 0.07 NP1 n.d. n.d.BM1Dimer 0.10 0.19 BM1EE 0.15 0.14 Unid RRT 0.05 n.d. 0.69

TABLE 8b.2 Related substances and assay of bendamustine HCl (residualcontent) in coated tablet Bendamustine HCl [% area] Storage Related T =2 T = 2 condition substances T = 0 months T = 0 months 40° C./75% RH HP10.19 0.16 99.46 99.36 (closed vials) HP2 n.d. 0.06 HP3 n.d. 0.05 NP1n.d. n.d. BM1Dimer 0.09 0.18 BM1EE 0.15 0.14 Unid RRT <0.05 <0.05 0.69

Example 9

TABLE 9a composition coated tablets Composition PF1 PF2 PF3 componentmg/tablet mg/tablet mg/tablet Bendamustine HCl 55.1 55.1 55.1 Anhydrousdextrose — — 205.8 Dextrose monohydrate 186.0 — — Trehalose — 66.0 —Sorbitol — — 43.9 Lactose DCL 21 68.2 185.7 — Avicel ® PH 112 18.7 —23.0 Crospovidone — 21.0 — Magnesium stearate 2.0 2.2 2.2 Opadry 8.0 8.08.0 Total 338.0 338.0 338.0

Manufacturing Method for Formulations PF1 for 600 Tablets:

33.06 g of bendamustine, 111.60 g of dextrose, 40.92 g of lactose, 11.22g of microcrystalline cellulose and 1.20 g of magnesium stearate wereweighed and transferred into a double polyethylene bag and mixed for 5minutes. Thereafter the powder blend was transferred to the hopper of aneccentric tabletting machine (Korsch EK0) and compressed into roundtablets having the following characteristics: mean value diameter: 10.0mm; mean value mass: 336.9 mg (begin)—335.98 (end); friability: 0.15%;mean hardness value: 69.25N (begin)—68.60N (end).

The tablet cores were subsequently coated in a coating pan (4M8 ForMatePanCoat) using a 9% Opadry® TM White aqueous suspension and dried. Themean mass of the tablets was 342.42 mg. Thereafter the tablets werepacked into amber glass bottles closed with screw plugs and stored at40° C./75% RH.

Manufacturing Method for Formulations PF2 for 600 Tablets:

33.06 g of bendamustine, 111.42 g of lactose, 39.60 g of trehalose,12.60 g of cross-linked polyvinylpyrrolidone and 1.32 g of magnesiumstearate were weighed and transferred into a double polyethylene bag andmixed for 5 minutes. Thereafter the powder blend was transferred to thehopper of an eccentric tabletting machine (Korsch EK0) and compressedinto round tablets having the following characteristics: mean valuediameter: 10.0 mm; mean value mass: 332.95 mg (begin)—332.12 (end);friability: 0.3%; mean hardness value: 65.9 N (begin)—59.0 N (end).

The tablet cores were subsequently coated in a coating pan (4M8 ForMatePanCoat) using a 9% Opadry® TM White aqueous suspension and dried. Themean mass of the tablets was 340.1 mg. Thereafter the tablets werepacked into amber glass bottles closed with screw plugs and stored at40° C./75% RH.

Manufacturing Method for Formulation PF3:

Sorbitol and anhydrous dextrose were weighed. 140.64 g of Sorbitol wasdissolved in 105.48 g of purified water and the solution obtained wassubsequently used to granulate 659.36 g of dextrose in a Fluid BedGranulator (4M8ForMate FluidBed). Thereafter the granulate was dried at60° C. and sieved through a 850 μm sieve.

33.06 g of bendamustine hydrochloride, 149.82 g of the sorbitol/dextrosegranulate, 13.8 g of microcrystalline cellulose and 1.32 g of magnesiumstearate were weighed into a double polyethylene bag and mixed for 5minutes. Thereafter the powder blend was transferred to the hopper of aneccentric tabletting machine (Korsch EK0O and compressed into roundtablets having a mean diameter of 10.0 mm. The tablets had a mean valuefor mass of 335.99 mg (begin)—339.50 mg (end); friability: 0%; meanhardness value: 125.60N (begin)—129.7N (end). The tablets were thensubjected to a conditioning process according to the following two steps(performed only on selected batches): placing them at 25° C./60% R.H.for two hours and subsequently at 40° C. for two hours.

The tablets were subsequently coated in a coating pan (4M8 ForMatePanCoat) using a 9% Opadry® TM White aqueous suspension. Mean mass ofthe tablets: 341.43 mg. Thereafter the tablets were packed into amberglass bottles closed with screw plugs and stored at 40° C./75% RH.

The amount of bendamustine hydrochloride and of related substances inthe stored film-coated tablets was measured with HPLC, as describedabove. The results are shown in Tables 9b.1-9b.3:

TABLE 9b.1 Related substances and assay of bendamustine HCl (residualcontent) in coated tablet (formulation 1; Opadry ®) PF1 Bendamustine HCl[% area] Storage Related T = 3 T = 3 condition substances T = 0 months T= 0 months 40° C./75% RH HP1 0.03 0.08 99.5 98.7 (closed vials) NP1 n.d.n.d. BM1Dimer 0.05 0.16 BM1EE 0.15 0.13 Individual 0.01 0.06 unknownimpurity

TABLE 9b.2 Related substances and assay of bendamustine HCl (residualcontent) in coated tablet (formulation 2; Opadry ®) PF2 Bendamustine HCl[% area] Storage Related T = 3 T = 3 condition substances T = 0 months T= 0 months 40° C./75% RH HP1 0.02 0.23 98.5 98.3 (closed vials) NP1 0.010.01 BM1Dimer 0.03 0.23 BM1EE 0.15 0.11 Individual 0.01 0.05 unknownimpurity

TABLE 9b.3 Related substances and assay of bendamustine HCl (residualcontent) in coated tablet (formulation 3; Opadry ®) PF3 Bendamustine HCl[% area] Storage Related T = 3 T = 3 condition substances T = 0 months T= 0 months 40° C./75% RH HP1 0.05 0.09 98.1 98.4 (closed vials) NP1 n.d.n.d. BM1Dimer 0.06 0.19 BM1EE 0.15 0.14 Individual 0.03 0.11 unknownimpurity

3. Dissolution Tests Example 10

Dissolution tests for the tablet formulations of Examples 2 and 3 werecarried out in artificial gastric fluid at T=0. The dissolution samplesare tested for assay by HPLC (column: Zorbax Bonus-RP, 5 μm; temperatureof column oven: 30° C.; temperature of autosampler: 5° C.; detector: 254nm). Artificial gastric fluid pH 1.5 was prepared by dissolving 2 g ofsodium chloride p.A. in 1000 ml of water and adjusting the pH to1.5±0.05 with 5 N hydrochloric acid. The dissolution test was conductedaccording to Chapter 2.9.3. of European Pharmacopoeia 6.0, usingApparatus 2 (Paddle-apparatus). The rotation speed of the paddle was 50rpm, the temperature was 37° C.±0.5° C., the amount of dissolutionmedium was 500 ml.

The results for the tablet formulations of Example 2 (tabletformulation 1) and Example 3 (tablet formulation 2) are shown in thefollowing Table 10a:

TABLE 10.a Tablet Tablet formulation 1 formulation 2 DissolutionDissolution: Single Mean Single Mean Dissolution value value value valueafter: [%] [%] [%] [%] 10 min 85.3 84 80.9 88 77.4 87.8 87.2 88.7 90.694.3 79.6 87.9 84.1 90.8 20 min 94.7 95 96.5 96 95.7 98.7 96.6 95.7 96.494.3 93.0 93.8 93.9 97.0 30 min 93.3 94 95.3 95 94.3 96.4 95.4 94.4 95.493.1 91.8 92.9 93.0 95.3

The results of the same dissolution tests carried out on the coatedtablet formulations of Example 6, Example 7 and Example 8 at T=0 areshown in the following Table 10b:

TABLE 10b Tablet Tablet Tablet formulation formulation formulationDissolution example 6 example 7 example 8 after Mean value Mean valueMean value 10 minutes 77 47 83 20 minutes 88 76 90 30 minutes 87 87 88

Corresponding dissolution data for the tablets of example 9 were:

Tablet Tablet Tablet formulation formulation formulation example 9 (PF1)example 9 (PF2) example 9 (PF3) Mean value after Mean value after Meanvalue after Dissolution 3 months at 3 months at 3 months storage atafter 40° C./75% RH 40° C./75% RH 40° C./75% RH 10 minutes 89.7 96.360.1 20 minutes 93.7 95.2 88.8 30 minutes 93.2 93.3 94.0

As may be taken from the above all tablet formulations of the inventionshow a fast dissolution of bendamustine. In particular the inventiveformulations show a dissolution profile of the bendamustine as definedhereinbefore.

4. In Vivo Tests

Animal Bioavailability Studies of Bendamustine were Performed in BeagleDogs: PK Study Outlines

Study 1

The objective was to determine the bioavailability of 1 dose (i.e. 50mg) of bendamustine in 3 tablet formulations (T1-3) and 1 capsuleformulation (C) with a total of 4 oral formulations: AUC and Cmax

Total number of animals required: 16

Basic Design:

Cross-over design, 8 animals per arm:

TABLE 11a Period 1 (single dose of tablet, or capsule, day 1): Dose #Number of Animal Group Treatment Dose route (mg) animals numbers 1Bendamustine Capsule 50 2 Male + 37, 39 2 Female 38, 40 2 BendamustineTablet T1 50 2 Male + 41, 43 2 Female 42, 44 3 Bendamustine Capsule 50 1Male + 45 1 Female 46 4 Bendamustine Tablet T2 50 2 Male + 47, 49 1Female 48 5 Bendamustine Tablet T3 50 1 Male + 51 2 Female 50, 52

One Week Wash-Out

TABLE 11b Period 2 (1 week after period 1, single dose of any of thefollowing, day 8): Dose # Number of Animal Group Treatment Dose route(mg) animals numbers 1 Bendamustine Tablet T1 50 2 Male + 37, 39 2Female 38, 40 2 Bendamustine Capsule 50 2 Male + 41, 43 2 Female 42, 443 Bendamustine Tablet T3 50 1 Male + 45 1 Female 46 4 BendamustineCapsule 50 2 Male + 47, 49 1 Female 48 5 Bendamustine Tablet T2 50 1Male + 51 2 Female 50, 52

One Week Wash-Out

TABLE 11c Period 3 (1 week after period 2, single dose of any of thefollowing, day 15): Dose Dose # Number of Animal Group Treatment route(mg) animals numbers 3 Bendamustine Tablet T2 50 1 Male + 45 1 Female 464 Bendamustine Tablet T3 50 2 Male + 47, 49 1 Female 48 5 BendamustineCapsule 50 1 Male + 51 2 Female 50, 52

Study 2

The objective was to determine the bioavailability of 1 dose (i.e. 50mg) of bendamustine in 1 tablet formulation T4, and 1 capsuleformulation (C) with a total of 3 oral formulations: AUC and Cmax

Total number of animals required: 16

Basic Design:

Cross-over design, 8 animals per arm:

TABLE 12a Period 1 (single dose of capsule, day 1): Dose # Number ofAnimal Group Treatment Dose (mg) animals numbers 1 Bendamustine Capsule50 4 Male + 4 Female 2 Bendamustine Capsule 50 4 Male + 4 Female

One Week Wash-Out

TABLE 12b Period 2 (1 week after period 1, single dose of either of thefollowing formulations, day 8): Dose # Number of Animal Group TreatmentDose route (mg) animals numbers 1 Bendamustine Formulation 50 4 Male + X4 Female 2 Bendamustine T4 50 4 Male + 4 Female

Example 11

The coated tablets of Example 9 (formulation 3, coated with Opadry®Tablets T4), containing 50 mg of bendamustine, were orally administeredto male and female dogs in comparison with the capsules of the referenceexample.

The mean plasma profiles vs. time for both the capsule formulation andthe coated tablet of Example 9 are shown in FIG. 1.

Example 12

The coated tablets of Examples 6, 7, or 8 (Tablets T1 to T3), containing50 mg of bendamustine, were orally administered to male and female dogsin comparison with the capsules of the reference example.

The mean plasma profiles vs. time of the capsule formulation and thecoated tablets of Examples 6 to 8 are shown in FIG. 1.

Experiments were conducted in order to:

assess which saccharides or saccharide mixtures are suitable to obtainchemically stable tablets, with fast dissolution profile and hardnessvalues suitable for coating;

evaluate the compatibility between API and excipients;

develop placebo and API-containing batches by investigating differentmanufacturing processes: dry granulation, direct compression and wetgranulation;

evaluate different bendamustine hydrochloride/saccharide weight ratios;

evaluate the impact of saccharide purity on the formation ofbendamustine hydrochloride purities;

investigate the influence of moisture content on the technologicalproperties and stability of the manufactured tablets;

manufacture tablets using the commercially available freeze driedbendamustine hydrochloride product (Ribomustin®) and to compare theproperties of these tablets with tablets produced using correspondingamounts of mannitol and bendamustine hydrochloride.

The following saccharidests were used for the manufacturing of tabletsin accordance with the invention, the tablets containing 50 mg ofbendamustine (55 mg as bendamustine hydrochloride)

TABLE 13 Product name/ Chemical name Manufacturer Class Dextroseanhydrous Dextrose anhydrous Monosaccharide C/Roquette Dextroseanhydrous Dextrose anhydrous ST Monosaccharide 0.5/Roquette Dextrosemonohydrate Dextrose monohydrate Monosaccharide G/Roquette Dextrosemonohydrate Dextrose monohydrate Monosaccharide M/Roquette Lactitolmonohydrate Lacty-M/Purac Biochem Disaccharide Lactitol MC/DaniscoTrehalose Treha 16400/Cargill Disaccharide Sorbitol NeosorbP60W/Roquette Monosaccharide Erythritol Zerose (TM) ErythritolMonosaccharide 16954/Cargill Maltose Monohydrate Sunmalt S/HayashibaraDisaccharide Mannitol Pearlitol 200 SD/Roquette Monosaccharide Lactoseanhydrous SuperTab 21 AN/DMV- Disaccharide Fonterra Excipients Lactosemonohydrate SuperTab 14 SD/DMV- Disaccharide Fonterra ExcipientsFructose Fructose MS/Galam Monosaccharide Maltitol SweetpearlDisaccharide P200/Roquette Xylitol Xylisorb 300/Roquette MonosaccharideSucrose Sucrose Comprizucker Disaccharide S/Suedzucher Sucrose SucroseRFS/Suedzucker Disaccharide Sucrose 97% + EV saccharide DC 3, 75Disaccharide Maltodextrin 3% MD/Vibar Nord SPA β-Cyclodextrin KleptoseDC/Roquette Cyclic eptasaccharide D-Raffinose n/a/Senn ChemicalsTrisaccharide Pentahydrate D-Melezitose n/a/Biosynth Trisaccharidemonohydrate Microcrystalline Avicel PH112/FMC Polysaccharide CelluloseBiopolymer Microcrystalline Avicel pH101/FMC Polysaccharide CelluloseBiopolymer

The quality of the batches made was assessed by observation of thephysical appearance, identification test (HPLC), dissolution test,content and related substances assay (HPLC), content uniformity test(HPLC), hardness test and water content (Karl Fischer method). Batcheswere submitted to accelerated stability studies packaged in amber glassbottles under the storage conditions detailed in the following table.For each manufactured API-containing batch some tablets were stored at5° C. as back-up samples.

In the following, the various excipients in relation to theirmanufacturing process were investigated. By using these excipientsseveral placebo manufacturing trials were carried out by dry granulationto obtain preliminary information about the manufacturing methodsuitable to obtain tablets with good quality.

Two types of disintegrants were used: microcrystalline cellulose(Avicel®PH 112), as a standard disintegrant, and cross-linkedpolyvinylpyrrolidone (Crospovidone®), used just for batch D001T/002. Thechoice of Crospovidone® for batch D001T/002 (filler: anhydrous lactose)was based on the similarity between this formulation and the prototypeformulation of example 9. Magnesium stearate was used as lubricant forall the batches produced. The dry granulation manufacturing process forplacebo trials consisted in the following steps:

-   -   1. The saccharide and a partial quantity of lubricant        (83.3%_(w/w) of the total amount) were accurately weighed and        then mixed in a polyethylene bag for 2 minutes.    -   2. The obtained mixture was compacted by using the tabletting        machine equipped with a 18 mm diameter punch.    -   3. The obtained slugs were sieved by using a 850 micron net.    -   4. The granulate was weighed and mixed with the disintegrant and        the remaining amount of the lubricant (16.7%_(w/w)) in a        polyethylene bag for 2 minutes and then tabletted by using a 10        mm diameter punch.

Table 14 and Table 15 summarize the composition of each Placeboformulation and the results of the analytical tests performed on boththe final mixtures and the tablets. In Table 16, observations madeduring the manufacturing process of placebo batches and/or during theiranalytical characterization are reported.

The analytical and physical test results carried out on placebo batchesD001T/001-D001T/002-D001T/004-D001T/013-D001T/015 showed that theseformulations are suitable to be manufactured by dry granulation andfurther investigated by the addition of the API. All the otherformulations are characterized by a powder difficult to compact and,when obtained, tablets with high friability.

Batch D001T/005 (filler: β-cyclodextrin) showed good behaviour in drymanufacturing process, high hardness, low friability but longdisintegration time. This formulation was further investigated byemploying a super disintegrant (Crospovidone®) and adding the API (seefollowing paragraph).

TABLE 14 Dry granulation - Placebo batches composition and analyticalresults (batches D001T/001 ÷ D001T/010). D001T/001 D001T/002 D001T/003D001T/004 D001T/005 Components (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w))(%_(w/w)) Placebo Batches manufactured by Dry Granulation LactoseMonohydrate 93.7 — — — — (SuperTab 14 SD) Lactose Anhydrous — 93.7 — — —(SuperTab 21 AN) Mannitol — — 93.7 — — (Pearlitol 200 SD) Sorbitol — — —93.7 — (Neosorb P60W) β-Cyclodextrin — — — — 93.7 (Kleptose DC) DextroseAnhydrous — — — — — (Dextrose Anhydrous C) Dextrose Monohydrate — — — —— (Dextrose Monohydrate G) D-Raffinose Pentahydrate — — — — — Trehalose— — — — — (Treha 16400) Erythritol (Zerose Erythritol 16954) Avicel PH112 5.7 — 5.7 5.7 5.7 Crospovidone — 5.7 — — — Magnesium Stearate 0.60.6 0.6 0.6 0.6 Results of analytical tests performed on final mixturesWater Content (%) 5.26 0.85 0.47 0.93 12.62 Flowability (seconds) 24.684.66 9.91 13.70 15.61 (Test performed according to (Nozzle 1, (Nozzle 3,(Nozzle 2, (Nozzle 1, (Nozzle 1, EP 6.0, par. 2.9.16) diameter =diameter = diameter = diameter = diameter = 10.0 mm) 25.0 mm) 15.0 mm)10.0 mm) 10.0 mm) Results of analytical tests performed on tabletsHardness (N) 70 99 86 148 127 Friability (%) 0.1 0.1 0.6 0.2 0.2 (Testperformed according to EP 6.0, par. 2.9.7) Mean Weight (mg/tablet) 360365 319 332 327 Disintegration (min · sec) 5′07″ 1′24″ 2′51″ 4′56″20′59″ (medium: buffer pH = 1.5) D001T/006 D001T/007 D001T/008 D001T/009D001T/010 Components (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w))Placebo Batches manufactured by Dry Granulation Lactose Monohydrate — —— — — (SuperTab 14 SD) Lactose Anhydrous — — — — — (SuperTab 21 AN)Mannitol — — — — — (Pearlitol 200 SD) Sorbitol — — — — — (Neosorb P60W)β-Cyclodextrin — — — — — (Kleptose DC) Dextrose Anhydrous 93.7 — — — —(Dextrose Anhydrous C) Dextrose Monohydrate — 93.7 — — — (DextroseMonohydrate G) D-Raffinose Pentahydrate — — 93.7 — — Trehalose — — —93.7 — (Treha 16400) Erythritol 99.5 (Zerose Erythritol 16954) Avicel PH112 5.7 5.7 5.7 5.7 — Crospovidone — — — — — Magnesium Stearate 0.6 0.60.6 0.6 0.5 Results of analytical tests performed on final mixturesWater Content (%) 0.47 8.47 14.59 9.36 N.A. Flowability (seconds) 27.7025.37 19.43 20.85 N.A. (Test performed according to (Nozzle 1, (Nozzle1, (Nozzle 1, (Nozzle 1, EP 6.0, par. 2.9.16) diameter = diameter =diameter = diameter = 10.0 mm) 10.0 mm) 10.0 mm) 10.0 mm) Results ofanalytical tests performed on tablets Hardness (N) N.A. 54 46 61 N.A.Friability (%) N.A. Test failure Test failure Test failure N.A. (Testperformed according to (39.4) (31.7) (44.9) EP 6.0, par. 2.9.7) MeanWeight (mg/tablet) N.A 365 337 335 N.A. Disintegration (min · sec) N.A.4′18″ 1′22″ 3′59″ N.A. (medium: buffer pH = 1.5) N.A. = not availablebecause the mixture is not suitable for tabletting process (see theobservations reported in table 5a)

TABLE 15 Dry granulation - Placebo batches composition and analyticalresults (batches D001T/011 ÷ D001T/025). D001T/ D001T/ D001T/ D001T/D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ 011 012 013 014 015 016 017018 019 025 Components (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w))(%_(w/w)) (%_(w/w) (%_(w/w) (%_(w/w) (%_(w/w) Placebo Batchesmanufactured by Dry Granulation Fructose 93.7 — — — — — — — — —(Fructose MS) Maltitol — 93.7 — — — — — — — — (Sweetpearl P200) MaltoseMonohydrate — — 93.7 — — — — — — — (Sunmalt S) Lactitol Monohydrate — —— 93.7 — — — — — — (Lacty M) Sucrose 97% + — — — — 93.7 — — — — —Maltodextrin 3% (EV Saccharide DC 3.75 MD) Sucrose — — — — — 99.5(Sucrose Comprizucker S) Sucrose — — — — — — 93.7 (Sucrose granular RFS)Xylitol — — — — — — — 99.5 (Xylisorb 300) β-Cyclodextrin — — — — — — — —93.8 (Kleptose DC) D-Melezitose 93.8 monohydrate Avicel PH 112 — 5.7 5.75.7 5.7 — 5.7 — — — Crospovidone 5.7 — — — — — — — 5.6 5.6 MagnesiumStearate 0.6 0.6 0.6 0.6 0.6 0.5 0.6 0.5 0.6 0.6 Results of analyticaltests performed on final mixtures Water Content (%) 0.42 0.38 5.71 5.450.78 N/A 0.32 N/A 12.30 Flowability (seconds) 19.09 22.54 16.57 23.415.37 N/A 17.38 N/A 18.64 Not flow (Test performed (Nozzle 1, (Nozzle 1,(Nozzle 1, (Nozzle 1, (Nozzle 3, (Nozzle 1, (Nozzle 1, (Nozzle 3,according to EP 6.0, diameter = diameter = diameter = diameter =diameter = diameter = diameter = diameter = par. 2.9.16) 10.0 mm) 10.0mm) 10.0 mm) 10.0 mm) 25.0 mm) 10.0 mm) 10.0 mm) 25.0 mm) Results ofanalytical tests performed on tablets Hardness (N) 17 33 130 69 69 N/A19 N/A 62 56 Friability (%) Test failure Test failure 0.2 Test failure0.4 N/A Test failure N/A 0.2 Test failure (Test performed (100.0)(100.0) (19.6) (78.0) (20.7) according to EP 6.0, par. 2.9.7) MeanWeight 380 388 328 338 349 N/A 383 N/A 338 328 (mg/tablet)Disintegration (min.) 5′52″ 6′40″ 5′09″ 6′32″ 5′47″ N/A 4′50″ N/A 4′01″3′30″ (medium: buffer pH = 1.5) N/A = not available because the mixtureis not suitable for tabletting process (see the observations reported intable 5a)

TABLE 16 Observations about manufacturing process, product technologicalproperties and analytical tests for each manufactured placebo batchPlacebo Dry Granulation process/ Batches obtained slugs TablettingProcess/obtained tablets Analytical tests on tablets D001T/001 Excellentslugs; easy to be sieved Easy to be tabletted; good tablets obtainedFast disintegration; low friability; medium hardness D001T/002 Goodslugs; easy to be sieved Easy to be tabletted; good tablets obtainedFast disintegration; low friability; high hardness D001T/003 Difficultto compact; high pressure Difficult to be tabletted; Fastdisintegration; high friability; needed to obtain slugs The powderadheres to punches; tabletting high hardness process was interruptedafter a few tablets D001T/004 Excellent slugs; easy to be sieved Fairlygood to be tabletted; good tablets Fast disintegration; low friability;obtained very high hardness D001T/005 Excellent slugs; easy to be sievedEasy to be tabletted; good tablets obtained Slow disintegration; lowfriability; high hardness D001T/006 Poor slugs, high friabilityImpossible to be tabletted; tabletting No tablets available foranalytical process interrupted testing D001T/007 Poor slugs, highfriability Good to be tabletted; fairly good tablets Fastdisintegration; very high friability, obtained above acceptance limit;medium hardness D001T/008 Poor slugs, high friability Good to betabletted; fairly good tablets Fast disintegration; very highfriability, obtained above acceptance limit; medium hardness D001T/009Good slugs; easy to be sieved Good to be tabletted; fairly good tabletsFast disintegration; very high friability, obtained above acceptancelimit; medium hardness D001T/010 Impossible to obtain slugs; not — —further processed D001T/011 Poor slugs, high friability Poor tabletsobtained (many tablets break Fast disintegration; very high friability,during tabletting process) above acceptance limit (all tablets brokenafter test); very low hardness D001T/012 Poor slugs, high friabilityPoor tablets obtained (many tablets break Fast disintegration; very highfriability, during tabletting process) above acceptance limit (alltablets broken after test); low hardness D001T/013 Excellent slugs; easyto be sieved Easy to be tabletted; good tablets obtained Fastdisintegration; low friability; very high hardness D001T/014 Good slugs;easy to be sieved Good to be tabletted; fairly good tablets Fastdisintegration; high friability, obtained above acceptance limit; mediumhardness D001T/015 Good slugs; easy to be sieved Good to be tabletted;good tablets obtained Fast disintegration; medium friability; mediumhardness D001T/016 Impossible to obtain slugs; not — — further processedD001T/017 Poor slugs, high friability Poor tablets obtained (manytablets break Fast disintegration; high friability, during tablettingprocess) above acceptance limit; low hardness D001T/018 Impossible toobtain slugs; not — — further processed D001T/019 Excellent slugs; easyto be sieved Easy to be tabletted; good tablets obtained Fastdisintegration; low friability; medium hardness D001T/025 Good slugs;easy to be sieved Good to be tabletted; fairly good tablets Fastdisintegration; very high friability, obtained above acceptance limit;medium hardnessBatches Manufactured by Dry Granulation with a 1:5 BendamustineHydrochloride/Saccharide Weight Ratio

The placebo formulations, evaluated as more suitable to manufacturetablets containing the active pharmaceutical ingredient (API) by drygranulation, were modified to include the API and two API/saccharideweight ratios were explored: 1:5 and 1:2.

In this paragraph, formulations with a 1:5 API/saccharide weight ratiosare described.

Two types of disintegrant were used: microcrystalline cellulose(Avicel®PH 112), as a standard disintegrant, and crosslinkedpolyvinylpyrrolidone (Crospovidone), used just for the batch D001T/022.Magnesium stearate was used as lubricant for all the batches produced.

The manufacturing process of the API-containing batches by drygranulation consisted in the following steps:

-   -   1. The saccharide, a partial quantity of lubricant (83.3%_(w/w)        of the total amount) and Bendamustine Hydrochloride were        accurately weighed and mixed in a double polyethylene bag for 5        minutes.    -   2. The powder blend was pressed by using the tabletting machine        equipped with 18 mm diameter punch.    -   3. To obtain a granulate, the produced slugs were sieved by        using a 850 micron net.    -   4. The granulate was weighed and mixed with the disintegrant and        the remaining amount of the lubricant (16.7%_(w/w)) in a double        polyethylene bag for 5 minutes.    -   5. The obtained mixture was tabletted by using a 10 mm diameter        punch.

Table 17 summarizes the composition of each API-containing formulationmanufactured and the results of the analytical tests performed on theAPI-containing final mixtures; table 18 summarizes the results of theanalytical tests performed on the obtained products.

TABLE 17 Dry granulation - API/Saccharide weight ratio 1:5.API-containing batches final mixture composition and analytical results.API-containing batches manufactured by Dry granulation API/Saccharideratio 1:5 D001T/020 D001T/021 D001T/022 D001T/023 D001T/024 Components(%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) Bendamustine HCL 15.316.6 16.6 16.6 15.7 Lactose Monohydrate 78.4 — — — — (SuperTab 14 SD)Sorbitol(Neosorb P60W) — 77.1 — — β-Cyclodeztrin — — 77.1 — (KleptoseDC) Maltose (Food grade) — — — 77.1 — (Sunmalt S) Sucrose 97% + — — — —78.0 Maltodextrin 3%) (EV Saccharide DC 3.75 MD) Avicel PH 112  5.7  5.7 5.7  5.7 Crospovidone —  5.7 — — Magnesium Stearate  0.6  0.6  0.6  0.6 0.6 Result of analytical tests performed on final mixture Flowability(seconds) 7.31 (Nozzle 2, 19.91 (Nozzle 1, 3.89 (Nozzle 3, 23.00 (Nozzle1, 7.99 (Nozzle 3, (Test performed diameter = diameter = diameter =diameter = diameter = according to EP 6.0, 15 mm) 10 mm) 25 mm) 10 mm)25 mm) par. 2.9.16)

TABLE 18 Dry granulation - API//Saccharide weight ratio 1:5.API-containing batches tablets analytical results. Specification Resultsof analytical tests performed on tablets Analytical Test LimitsD001T/020 D001T/021 D001T/022 D001T/023 D001T/024 IdentificationPositive Positive Positive Positive Positive Positive (HPLC) Mean WeightSpecific for 359.43 336.27 334.67 333.19 349.72 (mg/tablet) each Limits:Limits: Limits: Limits: Limits: formulation 342 ÷ 378 315.4 ÷ 348.6315.4 ÷ 348.6 315.4 ÷ 348.6 332.5 ÷ 367.5 Content Uniformity CompliesComplies Complies Complies Complies Complies (Test performed accordingto RSD 1.26 RSD 1.42 RSD 0.84 RSD 0.58 RSD 2.02 EP 6.0) Assay (%)95.0%-105.0% 96.9 93.5 97.4 93.8 97.7 (HPLC) Related substances (%)(HPLC) HPI ≦0.5% 0.12 0.10 0.08 0.11 0.14 BM1 Dimer ≦0.2% 0.04 0.05 0.050.05 0.04 BM1EE ≦0.5% 0.14 0.13 0.13 0.13 0.15 NP1 ≦0.2% n.d. n.d. 0.01n.d. 0.1 Individual unknown impurity ≦0.1% 0.01 n.d. n.d. n.d. n.d.Total impurities ≦1.5% 0.31 0.28 0.27 0.29 0.34 Dissolution Test(Medium: buffer pH = 1.5) (% 10 min) 80% in 30 72.9 72.1 88.0 60.0 75.5(% 20 min) minutes 87.6 85.9 88.9 79.2 89.6 (% 30 min) 87.2 84.7 87.484.7 90.3 Moisture content (%) — 4.72 1.00 11.3 5.13 0.88 Hardness (N)≧40N 67 89 77 151 55 Friability (%) ≦1.0% 0.2 0.2 0.1 0.2 0.4 (Testperformed according to EP 6.0, par. 2.9.7)

The results of analytical tests performed both on final mixtures andfinished products were in most cases good, mainly for Content Uniformityand Purity. All API-containing batches showed satisfactory massuniformity, homogeneity of API content, and a low impurities content.The impurity profile of all formulations was in compliance with thespecifications of API (see specification limits in the tables), thus nodegradation occurs during manufacturing process.

Two API-containing batches showed low values in API assay; this resultcould be due to the small batch size and to the losses during themanufacturing process and the samples for IPCs on the final mixtures.

API-Containing Batches Manufactured by Dry Granulation with a 1:2API/Saccharide Weight Ratio

All the saccharide previously investigated by thy granulation tomanufacture tablets with a 1:5 API/Saccharide weight ratio were alsoevaluated at a ratio of 1:2.

For the manufacturing process see above. In this case, the obtainedmixture was tabletted by using a 8 mm diameter punch.

Two types of disintegrant were used: microcrystalline cellulose (Avicel®PH 112), as a standard disintegrant, and crosslinkedpolyvinylpyrrolidone (Ccrospovidone®′, used just for the batchD001T/105. For this batch we have explored the use of Avicel® PH 112 andof Crospovidone®. The Crospovidone® was chosen according to the previouscyclodextrin based formulation manufactured by dry granulation with a1:5 API/Saccharide (see previous results).

Table 19 and Table 20 summarize the composition of each API-containingformulation manufactured by dry granulation with an API/Saccharideweight ratio of 1:2 and the results of the analytical tests performed onboth, the final mixtures and the tablets. All API-containing batchesshowed suitable uniformity of mass, homogeneity of API content and lowimpurities content. Friability and hardness values are, in the most ofthe cases, in compliance with the specifications. In the case of batchesD001T/093, D001T/095 and D001T/096, the results of the dissolution testperformed on 6 tablets showed out of specifications values with a highRSD and the test was extended to a sample of 12 tablets.

Cyclodextrin based tablets show good properties with both disintegrants(Avicel® PH 112 and Crospovidone®).

TABLE 19 Dry granulation - API/Saccharide weight ratio 1:2.API-containing batches final mixture composition and analytical results.D001T/091 D001T/092 D001T/093 D001T/094 D001T/105 D001T/095 D001T/096Components (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w))(%_(w/w)) API-containing Batches manufactured by Dry GranulationAPI/SaccharideSaccharide ratio 1:2 Bendamustine HCl 31.1 31.1 31.1 31.131.1 31.1 31.1 Lactose 62.3 — — — — — Monohydrate (Supertab 14 SD)Lactose Anhydrous — 62.3 — — — — (Supertab 21 AN) Sorbitol — — 62.3 — —— (Neosorb P60W) β-Ciclodextrine — — — 62.3 62.3 — — (Kleptose DC)Sucrose97% + — — — — 62.3 — Maltodextrine3% (EV SaccharideSaccharide DC3.75 MD) Maltose (Food grade) — — — — — — 62.3 (Sunmalt S) Avicel PH 112 5.9  5.9  5.9  5.9  5.9  5.9 Crospovidone  5.9 Magnesium Stearate  0.7 0.7  0.7  0.7  0.7  0.7  0.7 Results of analytical tests performed onfinal mixtures Flowability (seconds) Not flow Not flow Not flow Not flowNot flow Not flow Not flow (Test performed (Nozzle 3, (Nozzle 3, (Nozzle3, (Nozzle 3, (Nozzle 3, (Nozzle 3, (Nozzle 3, according to EP 6.0,diameter = diameter = diameter = diameter = diameter = diameter =diameter = par. 2.9.16) 25.0 mm) 25.0 mm) 25.0 mm) 25.0 mm) 25.0 mm)25.0 mm) 25.0 mm)

TABLE 20 Dry granulation - API/Saccharide weight ratio 1:2.API-containing batches tablets analytical results. Specification Resultsof analytical tests performed on tablets Analytical Test LimitsD001T/091 D001T/092 D001T/093 Identification Positive Positive PositivePositive (HPLC) Mean Weight Specific for 175.01 178.85 176.90 RSD 2.3(mg/tablet) each Limits: Limits: Limits: formulation 168.2 ÷ 185.9 168.2÷ 185.9 168.2 ÷ 185.9 Content Uniformity Complies Complies CompliesComplies (Test performed according to RSD 2.43 RSD 2.41 RSD 3.34 EP 6.0)Assay (%) 95.0%-105.0% 96.0 96.8 96.6 (HPLC) Related substances (%)(HPLC) HP1 ≦0.50% 0.08 0.28 0.11 BM1 Dimer ≦0.20% 0.04 0.03 0.04 BM1EE≦0.50% 0.13 0.13 0.13 NP1 ≦0.20% 0.01 0.01 0.01 Individual unknownimpurity ≦0.10% n.d. 0.04 0.06 Total impurities ≦1.50% 0.27 0.54 0.36Total impurities after storage 0.29 0.31 0.31 at 40° C./75% RH for 3months Dissolution Test 80% in 30 min n = 12 (Medium: buffer pH = 1.5)(% 10 min) 49.2 84.7 36.2 (RSD 15.7) (% 20 min) 75.2 92.4 54.6 (RSD14.0) (% 30 min) 84.7 93.0 65.2 (RSD 10.4) Dissolution after storage at89 92 86 40° C./75% RH for 3 months Moisture content (%) — 4.02 0.620.70 Hardness (N) ≧40 N 95 49 118 Friability (%) ≦1.0% 0.5 0.7 0.3 (Testperformed according to EP 6.0) Results of analytical tests performed ontablets Analytical Test D001T/094 D001T/105 D001T/095 D001T/096Identification Positive Positive PositiTve Positive (HPLC) Mean Weight176.06 176.40 175.81 RSD 6.1 180.81 RSD 1.3 (mg/tablet) Limits: Limits:Limits: Limits: 168.2 ÷ 185.9 168.2 ÷ 185.9 168.2 ÷ 185.9 168.2 ÷ 18.9Content Uniformity Complies Complies Complies Complies (Test performedaccording to RSD 3.84 RSD 2.69 RSD 2.86 RSD 3.41 EP 6.0) Assay (%) 96.697.2 97.7 99.3 (HPLC) Related substances (%) (HPLC) HP1 0.11 0.19 0.120.08 BM1 Dimer 0.04 0.05 0.04 0.04 BM1EE 0.14 0.13 0.12 0.13 NP1 0.010.01 0.01 0.02 Individual unknown impurity 0.05 0.02 0.05 0.03 Totalimpurities 0.35 0.40 0.37 0.31 Total impurities after storage 0.32 0.350.35 at 40° C./75% RH for 3 months Dissolution Test n = 12 n = 12(Medium: buffer pH = 1.5) (% 10 min) 68.8 75.0 58.2 (RSD 24.3) 57.5 (RSD22.2) (% 20 min) 88.8 92.1 73.8 (RSD 19.1) 75.1 (RSD 17.5) (% 30 min)92.4 92.8 82.5 (RSD 20.5) 84.6 (RSD 19.7) Dissolution after storage at92 75 89 40° C./75% RH for 3 months Moisture content (%) 8.30 8.70 0.714.06 Hardness (N) 110 100 75 125 Friability (%) 0.5 0.5 1.5 0.1 (Testperformed according to EP 6.0) n.d. = not detectedAPI-Containing Batches Manufactured by Direct Compression with a 1:5API/SaccharideSaccharide Weight Ratio

The saccharides with suitable characteristics to be manufactured by drygranulation were also explored by using direct compression developingtablets with a 1:5 API/Saccharide ratio.

Two types of disintegrant were used: microcrystalline cellulose (Avicel®PH 112), as a standard disintegrant, and crosslinkedpolyvinylpyrrolidone (Crospovidone®), used just for batch D001T/029.

This manufacturing process consisted of the following steps:

-   -   1. Weighing the API and the excipients.    -   2. Transferring the raw materials in a double polyethylene bag        and mixing for about 5 minutes until an homogeneous powder blend        is obtained.    -   3. Transferring of the powder blend in the hopper of the        tabletting machine.    -   4. Compression of the powder blend using an eccentric tablet        machine equipped with a 10 mm diameter punch.

The characteristics of the API-containing batches manufactured by directcompression are presented in the following table.

TABLE 21 Direct Compression - API/Saccharide weight ratio 1:5.API-containing batches final mixture composition and analytical results.API-containing batches manufactured by Direct Compression D001T/026D001T/027 D001T/028 D001T/029 D001T/030 (%_(w/w)) (%_(w/w)) (%_(w/w))(%_(w/w)) (%_(w/w)) Bendamustine HCL 16.6 16.6 15.3 16.6 15.7 LactoseMonohydrate — — 78.4 — — (Supertab 14 SD) Sorbitol 77.1 — — — (NeosorbP60W) β-Cyclodextrin — — — 77.1 (Kleptose DC) Maltose (Food grade) 77.1— — — — (Sunmalt S) Sucrose 97% + — — — — 78.0 Maltodextrin 3% (EVSaccharide DC 3.75 MD) Avicel PH 112  5.7  5.7  5.7  5.7 Crospovidone ——  5.7 — Magnesium Stearate  0.6  0.6  0.6  0.6  0.6 Flowability(seconds) 4.78 (Nozzle 3, 4.01 (Nozzle 3, Not flow (Nozzle 3, Not flow(Nozzle 3, 4.12 (Nozzle 3, (Test performed diameter = diameter =diameter = diameter = diameter = according to EP 25 mm) 25 mm) 25 mm) 25mm) 25 mm) 6.0, par. 2.9.16)The obtained results of the analytical tests are listed in table 22.

TABLE 22 Direct Compression - API/Saccharide weight ratio 1:5.API-containing batches tablets analytical results. Specification Resultof analytical Tests performed on tablets Analytical Test LimitsD001T/026 D001T/027 D001T/028 D001T/029 D001T/030 IdentificationPositive Positive Positive Positive Positive Positive (HPLC) Mean WeightSpecific for 333.80 332.25 363.86 331.41 356.61 (mg/tablet) each Limits:Limits: Limits: Limits: Limits: formulation 315.4 ÷ 348.6 315.4 ÷ 348.6342.0 ÷ 378.0 315.4 ÷ 348.6 332.5 ÷ 367.5 Content Uniformity CompliesComplies Complies Complies Complies Not (Test performed according to EPRSD 3.51 RSD 3.60 RSD 0.88 RSD 1.57 Complies 6.0) RSD 10.84 Assay (%)95.0%-105.0% 94.5 97.2 100.8 100.1 99.6 (HPLC) Related substances (%)(HPLC) HP1 ≦0.5% 0.10 0.11 0.12 0.13 0.11 BM1 Dimer ≦0.2% 0.04 0.04 0.040.04 0.04 BM1EE ≦0.5% 0.13 0.15 0.14 0.14 0.14 NP1 ≦0.2% 0.01 0.01 0.020.01 0.02 Individual unknown impurity ≦0.1% 0.03 0.03 0.03 0.03 0.03Total impurities ≦1.5% 0.31 0.34 0.35 0.35 0.34 Dissolution Test(Medium: buffer pH = 1.5) (% 10 min) 80% in 30 45.5 71.5 54.7 83.3 73.5(% 20 min) minutes 69.7 89.7 88.6 89.5 90.9 (% 30 min) 83.3 89.3 91.191.5 91.3 Moisture content (%) — 5.04 0.71 4.40 11.26 0.83 Hardness (N)≧40 N 106 108 74 99 92 Friability (%) ≦1.0% 0.2 0.2 0.2 0.1 0.8 (Testperformed according to EP 6.0, par. 2.9.7)

As reported in the above table the API-containing tablets manufacturedby direct compression showed no critical differences from the onesproduced by dry granulation except for batch D001T/030 (filler: Sucrose97%+Maltodextrin 3%) that showed a non homogeneous API content and aslight increase in the value of friability.

Wet Granulation: Placebo Exploratory Trials

Based on the results obtained in the first and second part of theproject, the saccharides not suitable for dry granulation or directcompression were investigated by wet granulation.

The present approach to investigate the wet granulation technology isshown below.

Each saccharide was granulated according to the steps described in theflow-sheet of FIG. 2. At the end of each step the wet granulatedsaccharide was dried and a compression trial was performed to evaluateif the granulate was suitable for tabletting. Placebo batches weremanufactured only for the granulated saccharides with doubtful resultsof the compression test. The compositions and the relevant analyticalresults of the placebo trials are reported in Table 23.

Placebo batches were manufactured according to the following steps:

-   -   1. Wet granulation of the saccharide with water or sorbitol        solution using Fluid Bed or High Shear granulator (see above        Flow-sheet of wet granulation manufacturing trials, and table        23)    -   2. Drying of the wet granulated saccharide in the Fluid Bed        granulator or in oven.    -   3. Sieving the granulated saccharide by using 850 and 710 micron        nets.    -   4. Weighing of all components of the formulation and mixing in a        polyethylene bag for 2 minutes.    -   5. Compression of the powder blend using an eccentric tablet        machine equipped with a 10 mm diameter punch.

Avicel PH 112 and magnesium stearate were used as disintegrant and aslubricant, respectively, for all the batches produced.

TABLE 23 Wet granulation. Placebo batches composition and IPC results.D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/D001T/ 032 034 035 045 051 054 055 057 058 070 075 Components (%_(w/w))(%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w))(%_(w/w)) (%_(w/w)) (%_(w/w)) Placebo Batches manufactured by WetGranulation Dextrose 93.66 — — — — — — — — — Anhydrous (DextroseAnhydrous ST 0.5) Dextrose — 93.66 — — — — — — — — Monohydrate (DextroseMonohydrate) Mannitol — — 93.67 — — — — — — — (Pearlitol 200 SD)D-Melezitose — — — 93.55 — — — — — — monohydrate Maltitol — — — — 93.70(Sweetpearl P200) Trehalose — — — — — 93.72 (Food grade) (Treha 16400)D-Raffinose — — — — — — 93.72 Pentahydrate Erythritol — — — — — — —93.52 93.56 (Food grade) (Zerose Erythritol 16954) Fructose 93.66(Fructose MS) Xylitol 93.71 (Xyilisorb 300) Avicel PH 112 5.7 5.7 5.75.7 5.7 5.7 5.7 5.8 5.7 5.8 5.7 Magnesium 0.6 0.6 0.6 0.8 0.6 0.6 0.60.7 0.6 0.7 0.6 Stearate Equipment and Fluid Bed High Fluid Bed FluidBed Fluid Bed Fluid Bed Fluid Bed Fluid Bed High High High BinderSolution Water Shear Water Water Sorbitol Sorbitol Sorbitol SorbitolShear Shear Shear utilized for Water solution solution solution solutionWater Water Water saccharide granulation Percentage of n/a n/a n/a n/a1.2 1.1 1.1 1.1 N/A N/A N/A sorbitol in the granulate (%_(w/w))Percentage of 1.03 7.93 0.20 5.08 0.20 0.22 15.40 0.22 0.13 0.40 0.19water in the granulate (%_(w/w)) Results of analytical tests performedon tablets Appearance Complies Complies Complies Complies CompliesComplies Complies Complies Complies Not Not Complies Complies Hardness(N) 98 82 277 96 76 60 64 15 30 N/A N/AAPI-Containing Batches Manufactured by Wet Granulation with a 1:5API/Saccharide weight ratio

Manufacturing trials including a wet granulation process were carriedout on all saccharides that turned out to be not suitable for tabletmanufacturing by dry granulation or direct compression technologies.

The manufacturing process of these trials performed at laboratory scaleis summarized as follow:

-   -   1. Wet granulation of the saccharide with water or sorbitol        solution using Fluid Bed or High Shear granulator (see above        Flow-sheet of wet granulation manufacturing trials, and table        24)    -   2. Drying of the wet granulated saccharide in the Fluid Bed        granulator or in oven    -   3. Sieving by using 850 and 710 micron nets.    -   4. Weighing of the API and excipients and mixing in a double        polyethylene bag for 5 minutes.    -   5. Compression of the powder blend using an eccentric tablet        machine equipped with a 10 mm diameter punch.

Avicel PH 112 and magnesium stearate were used as disintegrant and aslubricant, respectively, for all the batches produced.

Table 24 and Table 25 list the composition of each API-containingformulation manufactured by wet granulation and the results of theanalytical tests performed on both, the final mixtures and the tablets.

The results of the analytical tests performed on the final mixtures andon the finished products are, in the most of the cases, in compliancewith the specifications. No degradation occurs during the manufacturingprocess.

Among the saccharides investigated, only Fructose MS (Galam) is notsuitable to be processed by wet granulation: the API-containing batchD001T/047 has a high friability and the batch D001T/082 shows friabilityand hardness values out of specifications.

The batches D001T/060, D001T/061, D001T/082, D001T/086 have low valuesin API assay and for the batches D001T/082 and D001T/086 the Uniformityof Content does not comply, though the granulate was sieved by using 850micron and 710 micron nets. This result is probably due to poor powdersmixing.

TABLE 24 Wet granulation - API/Saccharide weight ratio 1:5.API-containing batches final mixture composition and analytical results.D001T/033 D001T/036 D001T/037 D001T/040 D001T/047 D001T/059 Components(%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w))(*) (%_(w/w))API-containing Batches manufactured by Wet Granulation Bendamustine HCl15.7 15.7 15.7 15.7 14.9 15.7 Dextrose Anhydrous 78 — — — — — (AnhydrousDextrose ST 0.5) Dextrose Monohydrate G — 78 — — — — Mannitol — — 78 — —— (Pearlitol 200 SD) Lactitol Monohydrate — — — 78 — — (Lacty M) (Foodgrade) D-Melezitose monohydrate — — — — 78.8 — Maltitol — — — — — 78(Sweetpearl P200) Trehalose (Food grade) — — — — — — (Treha 16400)D-Raffinose Pentahydrate — — — — — — Erythritol (Food grade) — — — — — —(Zerose Erythritol 16954) Xylitol (Xyilisorb 300) (**) Fructose MS (**)Avicel PH 112 5.7 5.7 5.7 5.7 5.7 5.7 Magnesium Stearate 0.6 0.6 0.6 0.60.6 0.6 Equipment and Binder Fluid Bed High Shear Fluid Bed Fluid BedFluid Bed Fluid Bed Solution utilized Water Water Water Water WaterSorbitol for saccharide granulation solution Percentage of sorbitol inthe N/A N/A N/A N/A N/A 1.2 granulate (%_(w/w)) Results of analyticaltests performed on final mixtures Flowability (seconds) Not flow 18.9511.14 6.12 Not flow 5.12 (Test performed according to (Nozzle 3, (Nozzle1, (Nozzle 2, (Nozzle 2, (Nozzle 3, (Nozzle 2, EP 6.0, par. 2.9.16)diameter = diameter = diameter = diameter = diameter = diameter = 25.0mm) 10.0 mm) 15.0 mm) 15.0 mm) 25.0 mm) 15.0 mm) Observations onmanufactured tablets Appearance Complies Complies Complies CompliesComplies Complies Stability Study Suitable for Suitable for Suitable forSuitable for Suitable for Suitable for Stability Stability StabilityStability Stability Stability Study Study Study Study Study StudyD001T/060 D001T/061 D001T/082 D001T/086 D001T/087 Components (%_(w/w))(%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) API-containing Batchesmanufactured by Wet Granulation Bendamustine HCl 15.7 15.7 15.7 15.715.7 Dextrose Anhydrous — — — — (Anhydrous Dextrose ST 0.5) DextroseMonohydrate G — — — — Mannitol — — — — (Pearlitol 200 SD) LactitolMonohydrate — — — — (Lacty M) (Food grade) D-Melezitose monohydrate — —— — Maltitol (Sweetpearl P200) Trehalose (Food grade) 78 (Treha 16400)D-Raffinose Pentahydrate — 78 Erythritol (Food grade) — — 78 (ZeroseErythritol 16954) Xylitol 78 (Xyilisorb 300)(**) Fructose MS(**) 78Avicel PH 112 5.7 5.7 5.7 5.7 5.7 Magnesium Stearate 0.6 0.6 0.6 0.6 0.6Equipment and Binder Fluid Bed Fluid Bed High Shear High Shear HighShear Solution utilized Sorbitol Sorbitol Sorbitol Sorbitol Sorbitol forsaccharide granulation solution solution solution solution solutionPercentage of sorbitol in the 1.1 1.1 3.4 4.8 3.0 granulate (%_(w/w))Results of analytical tests performed on final mixtures Flowability(seconds) 6.46 5.35 Not flow 4.23 Not flow (Test performed according to(Nozzle 2, (Nozzle 2, (Nozzle 3, (Nozzle 2, (Nozzle 3, EP 6.0, par.2.9.16) diameter = diameter = diameter = diameter = diameter = 15.0 mm)15.0 mm) 25.0 mm) 15.0 mm) 25.0 mm) Observations on manufactured tabletsAppearance Complies Complies Complies Complies Not Complies (***)Stability Study Suitable for Suitable for Suitable for Suitable for NotSuitable Stability Stability Stability Stability for Stability StudyStudy Study Study Study (*) This batch contains an excess of A.P.I(5.9%); (**) It was not possible to investigate granulation step usingfluid bed with saccharide solution because these saccharides are notfluidized with air stream; (***) The final mixture is not suitable fortabletting

TABLE 25 Wet granulation - API/Saccharide weight ratio 1:5.API-containing batches tablets analytical results. Specification Resultsof analytical tests performed on tablets Analytical Test LimitsD001T/033 D001T/036 D001T/037 D001T/040 D001T/047 D001T/059 D001T/060D001T/061 D001T/082 D001T/086 D001T/087 Identification Positive PositivePositive Positive Positive Positive Positive Positive Positive PositivePositive N/A (HPLC) Mean Weight Specific for 348.52 351.56 354.06 351.98368.66 347.49 350.37 349.04 352.09 351.78 N/A (mg/tablet) each Limits:Limits: Limits: Limits: Limits: Limits: Limits: Limits: Limits: Limits:formulation 332.5 ÷ 367.5 332.5 ÷ 367.5 332.5 ÷ 367.5 332.5 ÷ 367.5351.5 ÷ 388.5 332.5 ÷ 367.5 332.5 ÷ 367.5 332.5 ÷ 367.5 332.5 ÷ 367.5332.5 ÷ 367.5 Content Complies Complies Complies Complies CompliesComplies Complies Complies Complies Not Not N/A Uniformity (Test RSD1.50 RSD 3.64 RSD 1.35 RSD 2.82 RSD 1.11 RSD 3.36 RSD 3.38 RSD 2.99complies complies performed RSD 31.06 RSD 8.39 according to EP 6.0)Assay (%) 95.0%-105.0% 101.0 98.2 98.4 98.6 101.2 96.7 91.3 92.7 90.694.0 N/A (HPLC) Related N/A substances (%) (HPLC) HP1 ≦0.5% 0.08 0.080.09 0.08 0.10 0.07 0.07 0.13 0.05 0.21 BM1 Dimer ≦0.2% 0.04 0.04 0.040.04 0.04 0.04 0.04 0.04 0.04 0.04 BM1EE ≦0.5% 0.13 0.13 0.13 0.14 0.160.14 0.13 0.13 0.12 0.15 NP1 ≦0.2% 0.01 0.01 0.01 0.01 0.01 0.01 0.010.01 0.01 0.02 Individual ≦0.1% 0.02 0.02 0.02 0.03 0.03 n.d. 0.01 0.020.02 0.04 unknown impurity Total impurities ≦1.5% 0.30 0.30 0.31 0.300.34 0.26 0.26 0.33 0.24 0.49 Dissolution Test (Medium: buffer pH = 1.5)(% 10 min) 80% in 30 91.9 73.9 97.1 77.0 80.9 77.5 86.3 71.4 90.5 87.2N/A (% 20 min) minutes 93.3 90.5 95.5 88.4 93.5 87.2 99.7 88.4 89.0 92.2(% 30 min) 91.8 89.5 93.7 87.8 92.3 86.9 99.9 87.9 87.2 89.2 Moisturecontent — 1.15 6.58 0.59 4.48 4.14 0.49 7.81 12.12 0.51 0.56 0.90 (%)Hardness (N) ≧40 N 68 66 140 46 73 81 48 71 26 56 N/A Friability (%)≦1.0% 0.5 0.5 0.2 0.6 Test failure 0.4 0.4 0.3 Test failure 0.3 N/A(Test performed (28.3) (74) according to EP 6.0, par. 2.9.7)API-Containing Batches Manufactured by Wet Granulation with a 1:2API/Saccharide Weight Ratio

All saccharides previously investigated by wet granulation tomanufacture tablets with a 1:5 API/Saccharide weight ratio were alsoevaluated at a ratio of 1:2.

The fructose was not evaluated at a ratio of 1:2 because the obtainedgranulate is not suitable for tabletting.

Avicel PH 112 and magnesium stearate were used as disintegrant and aslubricant, respectively, for all the batches produced.

To improve the uniformity of the API content, these API-containingbatches were manufactured by applying the following approach:

-   -   1. Wet granulation of the saccharide by using procedures        previously optimized    -   2. Preparation of the API-containing mixture    -   3. Dry granulation of the mixture (Slugs production→Slugs        sieving)    -   4. Tabletting of the obtained mixture by using a 8 mm diameter        punch.

For the step 3 (Dry granulation of the mixture) see above.

Table 26 and table 27 report the compositions and the analytical resultsof the API-containing batches manufactured by using wet granulatedsaccharides with a API/Saccharide weight ratio of 1:2. Friability is, inthe most of the cases, out of specifications. The API/Saccharide weightchange does not compromise the technological properties of the D001T/084batch (Filler: granulated mannitol).

TABLE 26 Wet granulation - A.P.I./Saccharide weight ratio 1:2.API-containing batches final mixture composition and analytical results.API-containing Batches manufactured by Wet Granulation API/Saccharideratio 1:2 D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/D001T/ 114 115 084 116 117 118 119 120 123 124 Components (%_(w/w))(%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w))(%_(w/w))(*) (%_(w/w)) Bendamustine HCl 31.1 31.1 31.1 31.1 31.1 31.131.1 31.1 31.1 31.1 Dextrose 62.3 Monohydrate G Dextrose Anhydrous —62.3 — — — — — — — — (Anhydrous Dextrose ST 0.5) Mannitol — — 62.3 — — —— — — — (Pearlitol 200 SD) D-Melezitose monohydrate — — — 62.3 — — — — —— Maltitol — — — — 62.3 — — — — — (Sweetpearl P200) Trehalose (Foodgrade) — — — — — 62.3 (Treha 16400) D-Raffinose Pentahydrate — — — — — —62.3 Erythritol (Food grade) — — — — — — — 62.3 (Zerose Erythritol16954) Lactitol monohydrate⁽*⁾ — — — — — — — — 62.3 Xylitol 62.3(Xyilisorb 300) (**) Avicel PH 112 5.9 5.9 5.9 5.9 5.9 5.9 5.9 5.9 5.95.9 Magnesium Stearate 0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7 0.7 Equipmentand Binder High Fluid Bed Fluid Bed Fluid Bed Fluid Bed Fluid Bed FluidBed High Fluid Bed High Solution utilized for Shear Water Water WaterSorbitol Sorbitol Sorbitol Shear Water Shear saccharide granulationWater solution solution solution Sorbitol Sorbitol solution solutionPercentage of sorbitol in the N/A N/A N/A N/A 1.2 1.1 1.1 3.4 N/A 4.8granulate (%_(w/w)) Results of analytical Tests performed on finalmixtures Flowability (seconds) Not flow Not flow Not flow Not flow Notflow Not flow Not flow Not flow Not flow Not flow (Test performed(Nozzle 3, (Nozzle 3, (Nozzle 3, (Nozzle 3, (Nozzle 3, (Nozzle 3,(Nozzle 3, (Nozzle 3, (Nozzle 3, (Nozzle 3, according to EP 6.0,diameter = diameter = diameter = diameter = diameter = diameter =diameter = diameter = diameter = diameter = par. 2.9.16) 25.0 mm) 25.0mm) 25.0 mm) 25.0 mm) 25.0 mm) 25.0 mm) 25.0 mm) 25.0 mm) 25.0 mm) 25.0mm) ⁽*⁾As the lactitol used to develop the previous formulation(API/Saccharide weight ratio 1:5) is no longer commercially available,this batch was manufactured by using lactitol purchased by newmanufacturer (Lactitol MC by Danisco).

TABLE 27 Wet granulation - A.P.I/Saccharide weight ratio 1:2.API-containing batches tablets analytical results. Specification Resultsof analytical tests performed on tablets Analytical Test LimitsD001T/114 D001T/115 D001T/084 D001T/116 D001T/117 IdentificationPositive Positive Positive Positive Positive Positive (HPLC) Mean WeightSpecific for 178.67 184.41 177.14 174.81 178.70 (mg/tablet) each Limits:Limits: Limits: Limits: Limits: formulation 168.2 ÷ 185.9 168.2 ÷ 185.9168.2 ÷ 185.9 168.2 ÷ 185.9 168.2 ÷ 185.9 Content Uniformity CompliesComplies Complies Complies Complies Complies (Test performed accordingto RSD 2.3 RSD 2.9 RSD 2.40 RSD 3.4 RSD 4.0 EP 6.0) Assay (%)95.0%-105.0% 96.9 103.5 98.8 94.8 98.5 (HPLC) Related substances (%)(HPLC) HP1 ≦0.50% 0.07 0.14 0.09 0.07 0.08 BM1 Dimer ≦0.20% 0.03 0.040.04 0.04 0.04 BM1EE ≦0.50% 0.14 0.16 0.13 0.13 0.15 NP1 ≦0.20% 0.010.01 0.01 0.01 0.01 Individual unknown impurity ≦0.10% n.d. n.d. n.d.0.01 0.02 Total impurities ≦1.50% 0.25 0.35 0.27 0.27 0.33 DissolutionTest (Medium: buffer pH = 1.5) (% 10 min) 80% in 30 min 71.7 85.3 94.465.6 49.8 (% 20 min) 93.5 91.7 93.8 83.6 69.2 (% 30 min) 94.5 91.8 92.688.8 88.8 Moisture content (%) — 5.4 1.1 1.5 3.2 0.5 Hardness (N) ≧40 N47 46 67 49 41 Friability (%)  ≦1.0% Test failure Test failure 0.4 Testfailure 1.2 (Test performed according to (9.0) (41.3) (60.0) EP 6.0)Results of analytical tests performed on tablets Analytical TestD001T/118 D001T/119 D001T/120 D001T/123 D001T/124 IdentificationPositive Positive Positive Positive Positive (HPLC) Mean Weight 179.86177.30 183.26 180.33 173.38 (mg/tablet) Limits: Limits: Limits: Limits:Limits: 168.2 ÷ 185.9 168.2 ÷ 185.9 168.2 ÷ 185.9 168.2 ÷ 185.9 168.2 ÷185.9 Content Uniformity Complies Complies Not Not Complies (Testperformed according to RSD 1.7 RSD 1.4 Complies Complies RSD 2.7 EP 6.0)RSD 7.4 RSD 11.3 Assay (%) 98.0 96.9 100.4 98.0 96.7 (HPLC) Relatedsubstances (%) (HPLC) HP1 0.06 0.11 0.09 0.05 0.06 BM1 Dimer 0.04 0.040.04 0.04 0.03 BM1EE 0.15 0.14 0.14 0.15 0.13 NP1 0.01 0.01 0.01 0.010.01 Individual unknown impurity 0.02 0.02 0.04 0.03 0.03 Totalimpurities 0.32 0.34 0.37 0.32 0.29 Dissolution Test (Medium: buffer pH= 1.5) (% 10 min) 48.2 68.2 81.9 51.4 58.2 (% 20 min) 74.5 92.5 84.471.9 80.4 (% 30 min) 84.8 92.5 84.5 82.2 86.3 Moisture content (%) 6.39.4 0.8 3.6 0.5 Hardness (N) 44 50 18 43 45 Friability (%) Test failureTest failure Test failure Test failure 0.8 (Test performed according to(41.1) (16.8) (97.4) (16.0) EP 6.0)

Effect of the API/Mannitol Weight Ratio

Mannitol based tablets were manufactured investigating the followingAPI/mannitol ratios: (1:0.01, 1:0.1, 1:0.5, 1:1.7, 1:4, 1:5, 1:6 and1:10). The formulation with a 1:5 API/mannitol weight ratio (standardformulation) was reported above.

For the production of these batches Avicel PH 112 and magnesium stearatewere used as disintegrant and as lubricant respectively. Regarding themanufacturing process, for the 1:1.7, 1:4, and 1:6 ratios, wetgranulated mannitol, Bendamustine Hydrochloride and excipients wereaccurately weighed and mixed in a double polyethylene bag for 5 minutes.For batch D001T/110 (1:10 ratio) a premix was performed. In this case,Bendamustine Hydrochloride was mixed, for 5 min, with half quantity ofthe excipients mixture. Then, the obtained mixture was added to theremaining quantity of the excipients and mixed for additional 5 minutes.The final mixture was tabletted using the tabletting machine equippedwith a suitable punch (8 mm diameter punch for 1:1, 1:1.7 and 1:2ratios, 10 mm in the case of 1:4 and 1:6 ratios, 12 mm for 1:7 ratio and14 mm for 1:10 ratio).

With regard to the 1:0.01, 1:0.1, 1:0.5 ratios, we have applied themanufacturing process reported above (wet granulation of the saccharideand subsequent dry granulation), to improve the API content uniformity.The obtained mixture was tabletted using a 6 mm diameter punch.

The following tables (Table 28 and Table 29) summarize the compositionsand the analytical results of the API-containing formulationsmanufactured to study the effects of the different API/Mannitol ratios.The batches D001T/111, D001T/083 and D001T/106 showed high friabilityand for the batches D001T/106, D001T/108 and D001T/109 the Uniformity ofContent did not comply deviating from data trends previously obtained.This result may be due to the fact that these batches were producedusing a new lot of Bendamustine HCl (Lot number: F08-05873) that mayhave different physical properties.

TABLE 28 Effect of the A.P.I/Mannitol weight ratio. API-containingbatches final mixture composition A.P.I./Mannitol Ratio Study D001T/D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/ D001T/113 112 111 083 106 084 108 037 109 085 110 (%_(w/w)) (%_(w/w))(%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w))(%_(w/w)) (%_(w/w)) A.P.I./ 1:0.01 (*) 1:0.1 (*) 1:05 (*) 1:1 1:1.7 1:21:4 1:5 1:6 1:7 1:10 Saccharide Ratio Bendamustine 55.1 55.1 55.1 44.134.4 31.1 18.7 15.7 13.4 11.9 8.6 HCl Mannitol 0.55 5.51 27.6 44.1 58.562.3 74.7 78.0 80.7 82.9 86.1 Granulated (Pearlitol 200 SD) Avicel PH11243.7 38.7 16.6 11.1 6.4 5.9 5.9 5.7 5.3 4.6 4.6 Magnesium 0.7 0.7 0.70.7 0.7 0.7 0.7 0.6 0.6 0.6 0.7 Stearate Batch number of F08-03755F08-03755 F08-03755 F08-03755 F08-05873 F08-03755 F8-05873 F08-03755F8-05873 F08-03755 F08-03755 Bendamustine HCl Results of analyticaltests performed on final mixtures Flowability Not flow Not flow Not flowNot flow Not flow Not flow 5.20 11.14 2.59 10.90 10.06 (seconds) (Nozzle3, (Nozzle 3, (Nozzle 3, (Nozzle 3, (Nozzle 3, (Nozzle 3, (Nozzle 3,(Nozzle 2, (Nozzle 3, (Nozzle 2, (Nozzle 2, (Test performed diameter =diameter = diameter = diameter = diameter = diameter = diameter =diameter = diameter = diameter = diameter = according to 25.0 mm) 25.0mm) 25.0 mm) 25.0 mm) 25.0 mm) 25.0 mm) 25.0 mm) 15.0 mm) 25.0 mm) 15.0mm) 15.0 mm) EP 6.0, par. 2.9.16) (*) Batches manufactured by using theexperimental approach reported above □ Standard formulation 1:5API/Saccharide weight ratio

TABLE 29 Effect of the A.P.I./Mannitol weight ratio study.API-containing batches tablets analytical results. Specification Resultsof analytical tests performed on tablets Analytical Test LimitsD001T/113 D001T/112 D001T/111 D001T/083 D001T/106 D001T/084Identification Positive Positive Positive Positive Positive PositivePositive (HPLC) Mean Weight Specific for 100.28 104.51 95.95 126.76163.85 177.14 (mg/tablet) each Limits: Limits: Limits: Limits: Limits:Limits: formulation 95 ÷ 105 95 ÷ 105 95 ÷ 105 118.75 ÷ 131.3 152.0 ÷168.0 168.2 ÷ 185.9 Content Uniformity Complies Complies CompliesComplies Complies Not Complies (Test performed according RSD 2.7 RSD 3.2RSD 3.7 RSD 2.77 Complies RSD 2.40 to EP 6.0) RSD 14.24 Assay (%)95.0%-105.0% 101.6 104.1 95.4 99.3 96.8 98.8 (HPLC) Related substances(%) (HPLC) HP1 ≦0.50% 0.07 0.11 0.06 0.09 0.09 0.09 BM1 Dimer ≦0.20%0.04 0.04 0.04 0.04 0.04 0.04 BM1EE ≦0.50% 0.15 0.16 0.15 0.14 0.12 0.13NP1 ≦0.20% 0.01 0.01 0.01 0.01 0.01 0.01 Individual unknown ≦0.10% n.d.n.d. n.d. 0.02 n.d. n.d. impurity Total impurities ≦1.50% 0.27 0.32 0.260.31 0.26 0.27 Dissolution Test (Medium: buffer pH = 1.5) (% 10 min) 80%in 30 min 71.8 81.3 86.7 80.6 75.7 94.4 (% 20 min) 77.5 88.0 92.1 83.181.8 93.8 (% 30 min) 80.0 87.5 95.2 81.3 84.1 92.6 Moisture content (%)— 2.3 2.1 1.1 0.8 0.5 1.5 Hardness (N) ≧40N 88 85 67 63 70 67 Friability(%) ≦1.0% 0.7 0.9 Test failure Test failure Test failure 0.4 (Testperformed according (6.1) (5.2) (18.8) to EP 6.0) Results of analyticaltests performed on tablets Analytical Test D001T/108 D0011/037 D001T/109D001T/085 D001T/110 Identification Positive Positive Positive PositivePositive (HPLC) Mean Weight 299.05 354.06 410.0 469.55 646.24(mg/tablet) Limits: Limits: Limits: 389.5 ÷ Limits: Limits: 280.3 ÷309.8 332.5 ÷ 367.5 430.5 441.8 ÷ 488.3 608 ÷ 672 Content Uniformity NotComplies Not Complies Complies (Test performed according Complies 1.35Complies RSD 2.79 RSD 2.6 to EP 6.0) RSD 8.63 RSD 8.33 Assay (%) 97.198.4 97.1 95.1 99.4 (HPLC) Related substances (%) (HPLC) HP1 0.09 0.090.06 0.12 0.07 BM1 Dimer 0.04 0.04 0.04 0.03 0.04 BM1EE 0.13 0.13 0.130.14 0.14 NP1 0.01 0.01 0.05 0.01 0.01 Individual unknown n.d. 0.02 n.d.0.03 n.d. impurity Total impurities 0.27 0.31 0.28 0.33 0.26 DissolutionTest (Medium: buffer pH = 1.5) (% 10 min) 90.1 97.1 91.4 96.7 97.7 (% 20min) 89.8 95.5 91.1 94.6 97.2 (% 30 min) 88.4 93.7 90.6 92.8 96.3Moisture content (%) 0.4 0.6 0.5 0.4 0.6 Hardness (N) 226 140 227 181 91Friability (%) 0.3 0.2 0.5 0.3 0.9 (Test performed according to EP 6.0)□ Standard formulation 1:5 API/Saccharide weight ratio

Saccharides Combination Study

Table 30 and Table 31 report the results concerning the saccharidecombination study.

The following combinations were investigated:

Monosaccharide/Disaccharide 1:1

⁽*⁾Mannitol (Pearlitol 200 SD)/Lactose Anhydrous (SuperTab 21 AN)

Sorbitol (Neosorb P60W)/Maltose (Sunmalt S)

Oligosaccharide/Monosaccharide 1:1

⁽*⁾D-Melezitose monohydrate/⁽*⁾Dextrose anhydrous ST 0.5

⁽*⁾Raffinose Pentahydrate granulated/⁽*⁾Mannitol granulated (Pearlitol200 SD)

Oligosaccharide/Disaccharide 1:1

⁽*⁾Raffinose Pentahydrate granulated/Lactose Monohydrate (Supertab 14SD)

β-Ciclodextrine (Kleptose DC)/Sucrose (EV Saccharide)

(*) These saccharides were granulated by wet granulation (see page 32)

The manufacturing process consisted in direct compression of theunprocessed or granulated saccharide.

By using Avicel PH 112 and magnesium stearate as disintegrant and aslubricant, respectively, these batches were manufactured performing thefollowing steps:

-   -   1. The saccharides (or the granulated saccharide), Bendamustine        Hydrochloride and excipients were accurately weighed and mixed        in a double polyethylene bag for 5 minutes.    -   2. The obtained mixture was tabletted by using a 10 mm diameter        punch.

TABLE 30 Saccharides Combination Study. API-containing batches finalmixture composition and analytical results. D001T/049 D001T/074D001T/100 D001T/101 D001T/102 D001T/103 Components (%_(w/w)) (%_(w/w))(%_(w/w)) (%_(w/w)) (%_(w/w)) (%_(w/w)) Saccharide Combination StudyAPI-containing Batches Bendamustine HCl 14.89 15.74 15.74 15.74 15.7415.74 Saccharide combination Oligosaccharide/Monosaccharide 1:1D-Melezitose monohydrate/Dextrose 78.81 — — — — — anhydrous ST 0.5Raffinose Pentahydrate/Mannitol — — — — 77.96 — (Pearlitol 200 SD)Saccharide combination Oligosaccharide/Disaccharide 1:1 RaffinosePentahydrate/Lactose — — — — — 77.96 Monohydrate (Supertab 14SD)β-Ciclodextrine (Kleptose DC)/Sucrose — — — 77.96 — — (EV Saccharide)Saccharide combination Monosaccharide/Disaccharide 1:1 Sorbitol (NeosorbP60)/Maltose — — 77.96 — — — (Sunmalt S) Mannitol (Pearlitol 200SD)/Anhydrous — 77.96 — — — Lactose (SuperTab 21 AN) Avicel PH 112  5.70 5.70  5.70  5.70  5.70  5.70 Magnesium Stearate  0.60  0.60  0.60  0.60 0.60  0.60 Results of analytical tests performed on final mixturesFlowability (seconds) Not flow Not flow 5.24 5.25 Not flow Not flow(Test performed according to EP 6.0, par. (Nozzle 3, (Nozzle 3, (Nozzle3, (Nozzle 3, (Nozzle 3, (Nozzle 3, 2.9.16) diameter = diameter =diameter = diameter = diameter = diameter = 25.0 mm) 25.0 mm) 25.0 mm)25.0 mm 25.0 mm 25.0 mm

TABLE 31 Saccharide Combination Study. API-containing batches tabletsanalytical results. Specification Results of analytical tests performedon tablets Analytical Test Limits D001T/049 D001T/074 D001T/100D001T/101 D001T/102 D001T/103 Identification (HPLC) Positive PositivePositive Positive Positive Positive Positive Mean Weight Specific for365.96 346.06 351.56 349.60 354.13 348.83 (mg/tablet) each 351.5 ÷ 388.5332.5 ÷ 367.5 332.5 ÷ 36.5 332.5 ÷ 367.5 332.5 ÷ 367.5 332.5 ÷ 367.5formulation Content Uniformity Complies Not Complies Not CompliesComplies Complies (Test performed according to EP complies RSD 1.41Complies RSD 1.51 RSD 4.73 RSD 1.46 6.0) RSD 5.55 RSD 4.50 Assay (%)95.0%-105.0% 96.7 95.1 97.5 97.6 97.9 98.9 (HPLC) Related substances (%)(HPLC) HP1 ≦0.50% 0.10 0.06 0.06 0.12 0.09 0.09 BM1 Dimer ≦0.20% 0.040.03 0.04 0.04 0.04 0.04 BM1EE ≦0.50% 0.15 0.13 0.12 0.12 0.12 0.12 NP1≦0.20% 0.01 0.01 0.01 0.01 0.01 0.01 Individual unknown impurity ≦0.10%0.03 n.d. 0.01 0.02 n.d. 0.02 Total impurities ≦1.50% 0.33 0.23 0.240.31 0.26 0.28 Dissolution Test (Medium: buffer pH = 1.5) (% 10 min) 80%in 30 min 76.3 73.1 62.9 65.4 89.6 55.7 (% 20 min) 93.8 97.4 86.1 89.491.1 87.6 (% 30 min) 92.7 99.2 91.7 93.4 89.8 88.9 Moisture content (%)— 2.70 0.60 2.77 5.66 5.38 8.12 Hardness (N) ≧40N 73 147 216 144 93 118Friability (%)  ≦1.0% 0.4 0.1 0.1 0.2 Test failure 0.2 (Test performedaccording to EP (26.4) 6.0)

In general, the tablets manufactured for the saccharides combinationstudies show good properties. However, the batch D001T/102 (RaffinosePentahydrate/Mannitol (Pearlitol 200 SD)), show high friability and thebatches D001T/100 and D001T/049 are non homogeneous in API content.

Example 14 Freeze Dried Bendamustine HCl (Ribomustin) and BendamustineHCl/Mannitol Tablets (Api/Saccharide Weight Ratio 1:1.2)

Tablets containing bendamustine hydrochloride/mannitol in a weight ratioof 1:1.2 were prepared by using either freeze dried material obtainedfrom the commercially available product for intravenous application(Ribomustin®) or using wet granulated mannitol and Bendamustine HCl.

The manufacturing processes were performed according to the followingexperimental operations: the freeze dried powder was removed from theRibomustin® vials and was sieved using a 850 micron net. The obtainedpowder and the lubricant (magnesium stearate) were accurately weighedand mixed in a polyethylene bag for 5 minutes. The mixture was slowlytransferred in the pressing chamber of the tabletting machine and wasmanually pressed by using an 8 mm diameter punch in order to obtainsmall slugs. The slugs were sieved using a 850 micron net and theobtained granulate was manually pressed using a 8 mm diameter punch.

Bendamustine HCl/mannitol tablets were manufactured applying the sameoperating procedures as described above in this example.

The composition of the formulations is reported in table 32.

TABLE 32 Ribomustin and bendamustine/mannitol tablets. API- containingbatches final mixture composition. Ribomustin and bendamustine/ mannitoltablets D001T/125 D001T/126 (%_(w/w)) (%_(w/w)) A.P.I./Saccharide 1:1.21:13 Ratio Ribomustin Freeze- 99.36 dried(*) Bendamustine HCl 45.16Mannitol Granulated 54.20 (Pearlitol 200 SD) Magnesium Stearate  0.64 0.64 Batch number of F08-03755 Bendamustine HCl Flowability (seconds)N/A N/A (Test performed according to EP 6.0, par. 2.9.16)(*)Corresponding to 45.16% of Bendamustine HCl and 54.20% of Mannitol

Table 33 reports the data concerning the comparison between the tabletsobtained using the freeze dried bendamustine hydrochloride/mannitolmixture and the non freeze-dried bendamustine hydrochloride/mannitolmixture.

TABLE 33 Ribomustin and bendamustine/mannitol tablets. API- containingbatches tablets analytical results. Results of analytical testsSpecification performed on tablets Analytical Test Limits D001T/125D001T/126 Identification Positive Positive Positive (HPLC) Mean WeightSpecific for 123.45 121.79 (mg/tablet) each Limits: Limits: formulation115.9 ÷ 128.1 115.9 ÷ 128.1 RSD 6.02 RSD 2.88 Content UniformityComplies Complies Complies (Test performed RSD 4.05 RSD 3.35 accordingto EP 6.0) Assay (%) (HPLC) 95.0%-105.0% 98.6 99.5 Related substances(%) (HPLC) HP1 ≦0.50% 1.03 0.08 BM1 Dimer ≦0.20% 0.19 0.04 BM1EE ≦0.50%0.19 0.14 NP1 ≦0.20% 0.01 0.01 Individual unknown ≦0.10% 0.03 n.d.impurity Total impurities ≦1.50% 1.50 0.27 Dissolution Test (Medium:buffer pH = 1.5) (% 10 min) 80% in 30 min 93.3 57.7 (% 20 min) 94.6 80.0(% 30 min) 93.0 89.9 Moisture content (%) — 1.61 0.21 Hardness (N) ≧40N61 44 Friability (%) ≦1.0%  N/A Test failure (Test performed (15.6)according to EP 6.0)

Taking as reference target the impurity profile of the BendamustineHydrochloride API (see specification limits in the table), batchD001T/125 showed an out of specification value for HP1 impurity. Theresults of the dissolution test highlight that, although after 10minutes the dissolution profile of the tablets, containing thefreeze-dried bendamustine hydrochloride/mannitol mixture is faster, forboth formulations, after 30 minutes the dissolution is in compliancewith the current specifications. The friability is out of specificationfor batch D001T/126, whereas the test was not performed for batchD001T/125 due to lack of sufficient amounts of material.

INDUSTRIAL APPLICABILITY

The compositions according to the present invention show manyadvantages. They can be easily used by the patient without assistance ofsupervisory medical staff. Hence the time-consuming trips to thehospital may become obsolete, thereby increasing the patient compliance.

Since the dosage forms are solid, they can be swallowed as such, whichmeans that the patient does not need to wait until dissolution of theactive ingredient has been achieved. Further due to the good stabilityof the dosage forms they can be easily stored at room temperature andwithout the need of any special storage conditions.

By using the dosage forms according to the present invention, aconsiderable reduction of the volume of the dosage form may be achieved.The reduced size is desirable both from a manufacturing and handlingstandpoint and patient compliance.

Pharmaceutical compositions show a high dissolution in vitro reducingthe degradation of bendamustine in vivo, thus resulting in an improvedbioavailability of the bendamustine in vivo.

1. A pharmaceutical composition in a solid dosage form suitable for oraladministration, the composition comprising bendamustine or apharmaceutically acceptable ester, salt or solvate thereof as an activeingredient, and at least one pharmaceutically acceptable excipient,which is a pharmaceutically acceptable saccharide selected from thegroup consisting of one or more of a monosaccharide, a disaccharide, anoligosaccharide, a cyclic oligosaccharide, a polysaccharide and asaccharide alcohol, wherein the ratio by weight of the active ingredientto the saccharide excipient(s) is in the range of 1:1-5.
 2. Thepharmaceutical composition of claim 1, wherein the ratio by weight ofthe active ingredient to the saccharide is 1:2-5.
 3. The pharmaceuticalcomposition according to claim 1 or 2, which is in the form of a tablet,a granulate, or a pill.
 4. The pharmaceutical composition according toany one of claims 1 to 3, wherein the tablet or tablet granules, thegranulate or the pill are provided with a coating.
 5. The pharmaceuticalcomposition according to any one of claims 1 to 4, wherein the activeingredient is bendamustine hydrochloride.
 6. The pharmaceuticalcomposition according to any one of 1 to 5, which comprises 10 to 1000mg of the active ingredient and 30 to 5000 mg of the saccharideexcipient.
 7. The pharmaceutical composition according to one of claims1 to 6, wherein the saccharide excipient is selected from mannitol,maltitol, erythritol, xylitol, lactose, sucrose, glucose, sorbitol,maltose, trehalose, lactitol, dextrose and fructose.
 8. Thepharmaceutical composition according to one of claims 1 to 6, whereinthe saccharide excipient is selected from dextrose anhydrous, dextrosemonohydrate, lactitol monohydrate, trehalose, sorbitol, erythritol,maltose monohydrate, mannitol, lactose anhydrous, lactose monohydrate,maltitol, xylitol, sucrose, sucrose 97%+maltodextrin 3%, β-cyclodextrin,D-raffinose pentahydrate, D-melezitose monohydrate and microcrystallinecellulose.
 9. The pharmaceutical composition according to any one ofclaims 1 to 8, which further comprises a pharmaceutically acceptablelubricant, filler and/or disintegrant.
 10. A pharmaceutical compositionaccording to claim 1, which shows a dissolution of the bendamustine ofat least 60% in 10 minutes, 70% in 20 minutes and 80% in 30 minutes, asmeasured with a paddle apparatus at 50 rpm according to the EuropeanPharmacopoeia in 500 ml of a dissolution medium at a pH of 1.5.
 11. Thepharmaceutical composition according to any one of claims 1 to 10 forthe treatment of a medical condition which is selected from chroniclymphocytic leukemia, acute lymphocytic leukaemia, chronic myelocyticleukaemia acute myelocytic leukaemia, Hodgkin's disease, non-Hodgkin'slymphoma, multiple myeloma, breast cancer, ovary cancer, small cell lungcancer, non-small cell lung cancer, and an autoimmune disease.
 12. Thepharmaceutical composition according to any one of claims 1 to 11, whichis to be administered in combination with at least one further activeagent, wherein said further active agent is given prior, concurrently,or subsequently to the administration of the pharmaceutical composition.13. The pharmaceutical composition according to claim 12, wherein thefurther active agent is an antibody specific for CD20, an anthracyclinderivative, a vinca alkaloid or a platin derivative.
 14. Thepharmaceutical composition according to claim 13, wherein the antibodyspecific for CD20 is rituximab, wherein the anthracyclin derivative isdoxorubicin or daunorubicin, wherein the vinca alkaloid is vincristineand wherein the platin derivative is cisplatin or carboplatin.
 15. Thepharmaceutical composition according to any one of claims 1 to 14, whichis to be administered in combination with at least one corticosteroid,wherein said corticosteroid is given prior, concurrently, orsubsequently to the administration of the pharmaceutical composition 16.The pharmaceutical composition according to claim 15, wherein thecorticosteroid is prednisone or prednisolone.